LncRNA SPRY4-IT1 promotes progression of osteosarcoma by regulating ZEB1 and ZEB2 expression through sponging of miR-101 activity

Long non-coding (lnc)RNA sprouty receptor tyrosine kinase signalling antagonist 4-intronic transcript 1 (SPRY4-IT1) has been demonstrated to serve a critical role in the tumorigenesis of osteosarcoma (OS); however, the specific underlying mechanism remains unclear. The aim of the present study was to examine the interactions between SPRY4-IT1 and its downstream effectors, to determine if any of the interactions contributed to SPRY4-IT1-mediated proliferation, migration and invasion in cancer cells. A signalling cascade which involved SPRY4-IT1, miR-101 and zinc finger E-box-binding homeoboxes (ZEBs) was examined in the present study. Intracellular SPRY4-IT1 and miR-101 expression levels were altered through transfection to assess their effect on proliferation, cell cycle progression, survival, migration and invasion. A dual-luciferase assay was utilized to determine the association between SPRY4-IT1/miR-101 and ZEBs/miR-101 and nude mouse xenograft experiments were performed to determine the effect of SPRY4-IT1 in vivo. The results indicated that the SPRY4-IT1 levels were negatively associated with miR-101 expression levels in OS cells, an association which was not observed in the normal osteoblast cells. SPRY4-IT1 knockdown or miR-101 overexpression reduced proliferation, cell cycle progression, survival, migration and invasion of MG-63 and U2OS cells. SPRY4-IT1 knockdown was accompanied by increased expression of miR-101 and E-cadherin levels, as well as decreased expression levels of ZEB1/2 and other epithelial-mesenchymal transition-associated proteins. Simultaneous knockdown of SPRY4-IT1 and inhibition of miR-101 partially reversed the anti-tumour effects of SPRY4-IT1 inhibition in vitro. Consistent with these findings, short hairpin RNA targeting SPRY4-IT1 also hindered xenograft tumour growth and altered the levels of miR-101, ZEB1/2 and E-cadherin in vivo. Dual-luciferase reporter assays demonstrated that SPRY4-IT1 may have regulated the expression of ZEB1 and ZEB2 by sponging miR-101. In conclusion, SPRY4-IT1 inhibition increased miR-101 levels, resulting in downregulation of ZEB1/2 expression and thus exerting anti-tumour effects in OS.