Background: Osteosarcoma (OS) is a common bone tumor among children, adolescents, and young adults. Long non-coding RNA (lncRNA) FEZF1 antisense RNA 1 (FEZF1-AS1) has been reported as an oncogene in diverse tumors including colorectal cancer, pancreatic cancer and hepatocellular carcinoma, as well as in osteosarcoma. This study focused on the functions and mechanism of lncRNA FEZF1-AS1 in osteosarcoma.
Methods: The levels of FEZF1-AS1, microRNA miR-144 and CXC motif chemokine receptor 4 (CXCR4) in OS tissues and cells (Saos-2 and HOS) were measured by quantitative real-time polymerase chain reaction (qRT-PCR) or Western blot assay. The interactions between miR-144 and FEZF1-AS1 or CXCR4 were predicted by DIANA tools online database. Then, the dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were conducted to validate the interactions. Moreover, the cell viability and apoptotic rate in transferred Saos-2 and HOS cells were assessed via 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and flow cytometry, respectively. The levels of glucose and lactate productions were measured by glucose uptake and lactate production assay. In addition, the protein levels of Warburg-effect-related protein hexokinase 2 (HK2) and apoptosis-related proteins Bcl-2 or Bax in transferred Saos-2 and HOS cells were detected via Western blot assay.
Results: The levels of FEZF1-AS1 and CXCR4 were strikingly up-regulated, and miR-144 was notably down-regulated in OS tissues and cells. DIANA tools online database exhibited that miR-144 was a direct target of FEZF1-AS1 and CXCR4 was a direct target of miR-144. Then the interactions were validated by dual-luciferase reporter assay and RIP assay. Functionally, FEZF1-AS1 silencing or miR-144 overexpression inhibited cell viability, the glucose and lactate productions and promoted cell apoptosis in Saos-2 and HOS cells. Furthermore, miR-144 inhibitor mitigated the inhibitory effects on cell viability, the glucose and lactate productions and the promoted effect on cell apoptosis rate in Saos-2 and HOS cells induced by FEZF1-AS1 depletion. Mechanistically, FEZF1-AS1 regulated CXCR4 in Saos-2 and HOS cells by sponging miR-144.
Conclusion: We verified that FEZF1-AS1, CXCR4 were up-regulated, and miR-144 was downregulated in OS tissues and cells. Furthermore, FEZF1-AS1 promoted cell proliferation, Warburg effect and suppressed cell apoptosis in osteosarcoma via miR-144/CXCR4 axis, this novel pathway may provide a basis for the further study of osteosarcoma.
Keywords: lncRNA FEZF1-AS1, miR-144, CXCR4, Warburg effect, osteosarcoma