Department of Thoracic Surgery, Changsha Central Hospital, Changsha, Hunan, China.
Cell Cycle. 2023 May;22(10):1182-1195. doi: 10.1080/15384101.2022.2071565. Epub 2023 Mar 2.
Previous study has demonstrated the high expression of circular RNA 3-oxoacid CoA-transferase 1 (circ-OXCT1) in lung adenocarcinoma tumor tissues. However, the role and possible mechanism of circ-OXCT1 in non-small cell lung cancer (NSCLC) progression was unclear.Quantitative real-time PCR (qRT-PCR), western blotting and immunohistochemistry (IHC) staining assay were performed to detect the expression of circ-OXCT1, microRNA-516b-5p (miR-516b-5p), solute carrier family 1 member 5 (SLC1A5) and other indicated protein markers. Cell proliferation was measured by Cell counting kit 8 (CCK8), colony formation and 5-Ethynyl-2'-deoxyuridine (EdU) assays. Flow cytometry was employed to detect the rate of apoptotic cells. Cell migration and invasion were measured using transwell assay. The relative glutamine uptake and α-ketoglutarate (α-KG) production was determined using commercial kits. Interaction between miR-516b-5p and circ-OXCT1 or SLC1A5 was predicted by bioinformatics analysis and confirmed via luciferase reporter and RNA immunoprecipitation (RIP) assays. assay was implemented to demonstrate the effect of circ-OXCT1 in tumor growth.Circ-OXCT1 and SLC1A5 were upregulated and miR-516b-5p was downregulated in NSCLC tissues and cells. Functional experiments revealed that circ-OXCT1 silencing suppressed cell proliferation, migration and invasion, but promoted cell apoptosis . Circ-OXCT1 knockdown repressed tumor formation . Besides, miR-516b-5p was a target of circ-OXCT1, and miR-516b-5p inhibitor could relieve circ-OXCT1 absence-mediated effects in NSCLC cells. SLC1A5 was identified as a target of miR-516b-5p. Circ-OXCT1 promoted SLC1A5 expression by target binding with miR-516b-5p.Circ-OXCT1 facilitated NSCLC progression via miR-516b-5p-dependent regulation of SLC1A5, which provided a possible circRNA-targeted therapy for NSCLC.
先前的研究表明，环状 RNA 3-氧代酸 CoA 转移酶 1（circ-OXCT1）在肺腺癌肿瘤组织中高表达。然而，circ-OXCT1 在非小细胞肺癌（NSCLC）进展中的作用和可能的机制尚不清楚。实时定量 PCR（qRT-PCR）、蛋白质印迹和免疫组织化学（IHC）染色检测 circ-OXCT1、微小 RNA-516b-5p（miR-516b-5p）、溶质载体家族 1 成员 5（SLC1A5）和其他指示蛋白标志物的表达。细胞增殖通过细胞计数试剂盒 8（CCK8）、集落形成和 5-乙炔基-2'-脱氧尿苷（EdU）检测来测量。流式细胞术用于检测凋亡细胞的比率。使用 Transwell 测定法测量细胞迁移和侵袭。使用商业试剂盒测定相对谷氨酰胺摄取和α-酮戊二酸（α-KG）生成。通过生物信息学分析预测 miR-516b-5p 与 circ-OXCT1 或 SLC1A5 之间的相互作用，并通过荧光素酶报告和 RNA 免疫沉淀（RIP）测定进行验证。进行了实验以证明 circ-OXCT1 在肿瘤生长中的作用。circ-OXCT1 和 SLC1A5 在 NSCLC 组织和细胞中上调，而 miR-516b-5p 下调。功能实验表明，circ-OXCT1 沉默抑制细胞增殖、迁移和侵袭，但促进细胞凋亡。circ-OXCT1 敲低抑制肿瘤形成。此外，miR-516b-5p 是 circ-OXCT1 的靶标，miR-516b-5p 抑制剂可以减轻 NSCLC 细胞中 circ-OXCT1 缺失介导的作用。SLC1A5 是 miR-516b-5p 的靶标。circ-OXCT1 通过与 miR-516b-5p 靶结合促进 SLC1A5 的表达。circ-OXCT1 通过 miR-516b-5p 依赖性调节 SLC1A5 促进 NSCLC 进展，为 NSCLC 提供了一种可能的环状 RNA 靶向治疗方法。
