Department of Orthopedics, Changzhou No.2 People's Hospital, the Affiliated Hospital of Nanjing Medical University, No.29, Xinglong Lane, Tianning District, Changzhou, Jiangsu, China.
J Orthop Surg Res. 2022 Sep 15;17(1):421. doi: 10.1186/s13018-022-03303-1.
Accumulating articles demonstrate that circular RNAs play pivotal functions in tumorigenesis. However, the working mechanism of circ_0136666 in osteosarcoma (OS) progression remains to be further clarified.
Real time-quantitative polymerase chain reaction and western blot assay were applied to determine RNA and protein expression, respectively. Cell proliferation was assessed by 5-Ethynyl-2'-deoxyuridine assay and colony formation assay. Transwell assays were carried out to assess cell migration and invasion abilities. Flow cytometry was performed to analyze cell apoptosis. Cell glycolysis was evaluated by analyzing the uptake of glucose and the production of lactate using the corresponding kits. Dual-luciferase reporter assay and biotinylated RNA-pull down assay were performed to confirm the target interaction between microRNA-1244 (miR-1244) and circ_0136666 or centrosomal protein 55 (CEP55). Xenograft tumor model was utilized to explore the role of circ_0136666 in tumor growth in vivo.
Circ_0136666 expression was prominently elevated in OS tissues and cell lines. Circ_0136666 absence restrained the proliferation, migration, invasion and glycolytic metabolism and promoted the apoptosis of OS cells. Circ_0136666 negatively regulated miR-1244 expression by binding to it in OS cells. MiR-1244 overexpression suppressed the malignant behaviors of OS cells. CEP55 was a target of miR-1244 in OS cells. Circ_0136666 positively regulated CEP55 expression partly by sequestering miR-1244 in OS cells. CEP55 overexpression largely reversed circ_0136666 silencing-mediated influences in OS cells. Circ_0136666 silencing significantly suppressed tumor growth in vivo.
Circ_0136666 silencing inhibited OS progression partly by targeting miR-1244/CEP55 signaling. Silencing circ_0136666 and CEP55 or restoring miR-1244 level might be a potential therapeutic strategy for OS.
越来越多的文章表明，环状 RNA 在肿瘤发生中发挥着关键作用。然而，circ_0136666 在骨肉瘤（OS）进展中的作用机制仍有待进一步阐明。
实时定量聚合酶链反应和 Western blot 检测分别用于测定 RNA 和蛋白质表达。5-乙炔基-2'-脱氧尿苷检测和集落形成检测用于评估细胞增殖。Transwell 检测用于评估细胞迁移和侵袭能力。流式细胞术用于分析细胞凋亡。通过相应试剂盒分析葡萄糖摄取和乳酸生成来评估细胞糖酵解。双荧光素酶报告基因检测和生物素化 RNA 下拉检测用于证实 microRNA-1244（miR-1244）与 circ_0136666 或中心体蛋白 55（CEP55）之间的靶相互作用。利用异种移植肿瘤模型探讨 circ_0136666 在体内肿瘤生长中的作用。
circ_0136666 在 OS 组织和细胞系中显著上调。circ_0136666 缺失抑制 OS 细胞的增殖、迁移、侵袭和糖酵解代谢，并促进 OS 细胞凋亡。circ_0136666 在 OS 细胞中通过与 miR-1244 结合负调控 miR-1244 的表达。miR-1244 过表达抑制 OS 细胞的恶性行为。CEP55 是 OS 细胞中 miR-1244 的靶基因。circ_0136666 通过在 OS 细胞中与 miR-1244 结合负调控 CEP55 的表达。在 OS 细胞中过表达 CEP55 可显著逆转 circ_0136666 沉默介导的影响。circ_0136666 沉默显著抑制体内肿瘤生长。
circ_0136666 沉默通过靶向 miR-1244/CEP55 信号通路抑制 OS 进展。沉默 circ_0136666 和 CEP55 或恢复 miR-1244 水平可能是 OS 的一种潜在治疗策略。
