Jin Tao, Liu Mingfa, Liu Yan, Li Yuanzhi, Xu Zhennan, He Haoqi, Liu Jie, Zhang Yuxuan, Ke Yiquan
The National Key Clinical Specialty, Guangdong Provincial Key Laboratory on Brain Function Repair and Regeneration, Department of Neurosurgery, The Engineering Technology Research Center of Education Ministry of China, Zhujiang Hospital, Southern Medical University, Guangzhou, China.
Department of Neurosurgery, Shantou Central Hospital, Affiliated Shantou Hospital of Sun Yat-sen University, Shantou, China.
Front Oncol. 2020 Feb 21;10:170. doi: 10.3389/fonc.2020.00170. eCollection 2020.
Glioma is the most common malignant tumor of the central nervous system, and often displays invasive growth. Recently, circular RNA (circRNA), which is a novel non-coding type of RNA, has been shown to play a vital role in glioma tumorigenesis. However, the functions and mechanism of lipocalin-2 (Lcn2)-derived circular RNA (hsa_circ_0088732) in glioma progression remain unclear. We evaluated hsa_circ_0088732 expression by fluorescence hybridization (FISH), Sanger sequencing, and PCR assays. Cell apoptosis was evaluated by flow cytometry and Hoechst 33258 staining. Transwell migration and invasion assays were performed to measure cell metastasis and viability. In addition, the target miRNA of hsa_circ_0088732 and the target gene of miR-661 were predicted by a bioinformatics analysis, and the interactions were verified by dual-luciferase reporter assays. RAB3D expression was analyzed by an immunochemistry assay, and E-cadherin, N-cadherin, and vimentin protein expression were examined by western blot assays. A mouse xenograft model was developed and used to analyze the effects of hsa_circ_0088732 on glioma growth . We verified that hsa_circ_0088732 is circular and highly expressed in glioma tissues. Knockdown of hsa_circ_0088732 induced glioma cell apoptosis and inhibited glioma cell migration, invasion, and epithelial-mesenchymal transition (EMT). We found that hsa_circ_0088732 negatively regulated miR-661 by targeting miR-661, and was a target gene of miR-661. In addition, inhibition of miR-661 promoted glioma cell metastasis and suppressed cell apoptosis. Knockdown of induced cell apoptosis and suppressed cell metastasis. Moreover, hsa_circ_0088732 accelerated glioma progression through its effects on the miR-661/RAB3D axis. Finally, results from a mouse xenograft model confirmed that knockdown of hsa_circ_0088732 induced miR-661 expression, resulting in suppression of RAB3D expression and inhibition of tumor growth . We demonstrated that hsa_circ_0088732 facilitated glioma progression by sponging miR-661 to increase RAB3D expression. This study provides a theoretical basis for understanding the development and occurrence of glioma, as well as for the development of targeted drugs.
胶质瘤是中枢神经系统最常见的恶性肿瘤，常呈浸润性生长。近年来，环状RNA（circRNA）作为一种新型的非编码RNA，已被证明在胶质瘤的发生发展中起着至关重要的作用。然而，脂钙蛋白-2（Lcn2）衍生的环状RNA（hsa_circ_0088732）在胶质瘤进展中的功能和机制仍不清楚。我们通过荧光杂交（FISH）、桑格测序和PCR检测评估了hsa_circ_0088732的表达。通过流式细胞术和Hoechst 33258染色评估细胞凋亡。进行Transwell迁移和侵袭实验以检测细胞转移和活力。此外，通过生物信息学分析预测hsa_circ_0088732的靶标miRNA和miR-661的靶标基因，并通过双荧光素酶报告基因实验验证相互作用。通过免疫组织化学检测分析RAB3D表达，通过蛋白质印迹实验检测E-钙黏蛋白、N-钙黏蛋白和波形蛋白的蛋白表达。建立小鼠异种移植模型并用于分析hsa_circ_0088732对胶质瘤生长的影响。我们证实hsa_circ_0088732呈环状且在胶质瘤组织中高表达。敲低hsa_circ_0088732可诱导胶质瘤细胞凋亡并抑制胶质瘤细胞迁移、侵袭和上皮-间质转化（EMT）。我们发现hsa_circ_0088732通过靶向miR-661负向调节miR-661，且是miR-661的靶标基因。此外，抑制miR-661可促进胶质瘤细胞转移并抑制细胞凋亡。敲低诱导细胞凋亡并抑制细胞转移。此外，hsa_circ_0088732通过其对miR-661/RAB3D轴的影响加速胶质瘤进展。最后，小鼠异种移植模型的结果证实敲低hsa_circ_0088732可诱导miR-661表达，导致RAB3D表达受抑并抑制肿瘤生长。我们证明hsa_circ_0088732通过吸附miR-661增加RAB3D表达促进胶质瘤进展。本研究为理解胶质瘤的发生发展以及靶向药物的开发提供了理论依据。
