Zheng Shengnai, Qian Zhanyang, Jiang Fan, Ge Dawei, Tang Jian, Chen Hongtao, Yang Jin, Yao Yilun, Yan Junwei, Zhao Lei, Li Haijun, Yang Lei
Department of Orthopedic Surgery, Nanjing First Hospital, Nanjing Medical University Nanjing 210006, Jiangsu, PR China.
Department of Orthopedics, The First Affiliated Hospital of Nanjing Medical University Nanjing 210029, Jiangsu, PR China.
Am J Transl Res. 2019 Jul 15;11(7):4126-4138. eCollection 2019.
We aimed to investigate the biological functions of circLRP6 in the progression of osteosarcoma. CircLRP6 level in OS was detected by quantitative real-time polymerase chain reaction. Correlation between circLRP6 level with survival of OS patients was evaluated. Cell counting kit-8 and Transwell assay were conducted to detect proliferative, migratory and invasive capacities of OS cells. Cell cycle and apoptosis in OS cells influenced by circLRP6 were evaluated by flow cytometry. RNA immunoprecipitation was conducted to verify the binding relationship between circLRP6 with LSD1 and EZH2. Finally, the interaction between LSD1, EZH2 and promoter regions of KLF2, APC was clarified by chromatin immunoprecipitation. CircLRP6 level markedly increased in OS tissues. Besides, OS patients with high expression of circLRP6 showed shorter disease-free survival and over-all survival than those with low expression. CircLRP6 knockdown suppressed proliferative, migratory and invasive rates of OS cells. Moreover, circLRP6 knockdown induced apoptosis and arrested cell cycle in G0/G1 phase. The interaction between circLRP6 with LSD1 and EZH2 mediates their binding to the promoter regions of KLF2 and APC. Knockdown of circLRP6 weakened the binding abilities of LSD1, EZH2 to KLF2, APC. APC overexpression inhibited proliferation, induced apoptosis and arrested cell cycle. Moreover, the tumor-suppressor effect of downregulated circLRP6 on OS could be reversed by APC knockdown. Collectively, circLRP6 was highly expressed in OS and served as an oncogene by binding to LSD1 and EZH2 to inhibit expressions of KLF2 and APC.
我们旨在研究环状LRP6在骨肉瘤进展中的生物学功能。通过定量实时聚合酶链反应检测骨肉瘤中环状LRP6的水平。评估环状LRP6水平与骨肉瘤患者生存率之间的相关性。采用细胞计数试剂盒-8和Transwell实验检测骨肉瘤细胞的增殖、迁移和侵袭能力。通过流式细胞术评估环状LRP6对骨肉瘤细胞周期和凋亡的影响。进行RNA免疫沉淀以验证环状LRP6与LSD1和EZH2之间的结合关系。最后，通过染色质免疫沉淀阐明LSD1、EZH2与KLF2、APC启动子区域之间的相互作用。环状LRP6水平在骨肉瘤组织中显著升高。此外，环状LRP6高表达的骨肉瘤患者的无病生存期和总生存期均短于低表达患者。敲低环状LRP6可抑制骨肉瘤细胞的增殖、迁移和侵袭率。此外，敲低环状LRP6可诱导细胞凋亡并使细胞周期停滞在G0/G1期。环状LRP6与LSD1和EZH2之间的相互作用介导它们与KLF2和APC启动子区域的结合。敲低环状LRP6会削弱LSD1、EZH2与KLF2、APC的结合能力。APC过表达可抑制增殖、诱导凋亡并使细胞周期停滞。此外，敲低APC可逆转下调环状LRP6对骨肉瘤的肿瘤抑制作用。总的来说，环状LRP6在骨肉瘤中高表达，并通过与LSD1和EZH2结合来抑制KLF2和APC的表达而发挥癌基因作用。
