RESUMEN
Previous studies have supported a tumor-suppressive role of semaphorin 3A (SEMA3A) in several tumors including oral squamous cell carcinoma (OSCC). However, in-depth characterization of the role of SEMA3A in OSCC and the underlying molecular mechanisms is lacking. Gene and protein expressions were detected using quantitative real-time PCR, western blot assay, and immunohistochemistry. OSCC cell metastasis was evaluated using Transwell and angiogenesis of human umbilical vein endothelial cells (HUVECs) was determined using tube formation assay. The interactions among molecules were predicted using bioinformatics analysis and validated using luciferase activity experiment and RNA immunoprecipitation assay. Pulmonary metastasis was evaluated using hematoxylin and eosin staining after constructing a lung metastasis tumor model in mice. SEMA3A expression was decreased in OSCC cells and its overexpression led to suppression of epithelial-mesenchymal transition (EMT), migration, and invasion of OSCC cells and angiogenesis of HUVECs. miR-32-5p was identified as an upstream molecule of SEMA3A and long non-coding RNA NR2F2 antisense RNA 1 (NR2F2-AS1) was validated as an upstream gene of miR-32-5p. Further experiments revealed that the inhibitory effects of NR2F2-AS1 overexpression on EMT, migration, invasion of OSCC cells, and angiogenesis of HUVECs as well as tumor growth and metastasis in mice were mediated via the miR-32-5p/SEMA3A axis. To conclude, NR2F2-AS1 may attenuate OSCC cell metastasis and angiogenesis of HUVECs and suppress tumor growth and metastasis in mice via the miR-32-5p/SEMA3A axis.
RESUMEN
Oral squamous cell carcinoma (OSCC) is one of the most common carcinomas of the oral cavity. However, the regulatory mechanisms on miR-32-5p remain poorly understood in OSCC. The expression of miR-32-5p, Krüppel-like factor 2 (KLF2), C-X-C motif chemokine receptor 4 (CXCR4), and epithelial-to-mesenchymal transition (EMT)-related proteins (E-cadherin, Vimentin, N-cadherin, and Snail) were evaluated were assessed using RT-qPCR and Western blot. 3-(4, 5-Dimethylthiazolyl2)-2, 5-diphenyltetrazolium bromide assay, wound healing assay, and transwell assay were employed to detect cell proliferation, migration, and invasion of OSCC cells. Finally, dual-luciferase reporter assay was performed to verify the binding relationship between KLF2 and miR-32-5p. MiR-32-5p was highly expressed while KLF2 was lowly expressed in OSCC cells, and miR-32-5p knockdown or KLF2 overexpression could markedly reduce cell proliferation, migration, invasion, and EMT of OSCC cells. What is more, KLF2 was the target of miR-32-5p, and knockdown of KLF2 abolished the inhibitory effect of miR-32-5p inhibitor on progression of OSCC. Finally, CXCR4 expression was negatively regulated by KLF2, and inhibition of CXCR4 obviously alleviated the biological effects of si-KLF2 on the progression of OSCC. MiR-32-5p could enhance cell proliferation, migration, invasion, and EMT of OSCC cells, and the discovery of miR-32-5p/KLF2/CXCR4 axis might provide potential therapeutic targets for OSCC.