Fig. 6

CircRYK enhances GBM cell EMT and GSC maintenance through the miR-330-5p/VLDLR pathway. (A-B) qRT‒PCR tests were applied to assess the expression levels of VLDLR in GBM cells after the suppression of miR-330-5p alone or in combination with circRYK. (C) In the designated cells, VLDLR was examined using Western blotting, with β-actin serving as a control. (D-E) For analysis of the migration and invasion of cells using the Transwell method, GBM cells were genetically modified with shcircRYK, antagomir-330-5p, or a combination of antagomir-330-5p and shcircRYK. (F-G) For analysis of the invasion of the cells by three-dimensional spheroid assays, shcircRYK, antagomir-330-5p, or antagomir-330-5p and shcircRYK were transfected into U87-GFP and pGBM-1-GFP cells. (H) The wound healing test was utilized to examine the migration of GBM cells that were transfected with shcircRYK, antagomir-330-5p, or both antagomir-330-5p and shcircRYK. (I) Western blotting was utilized to investigate the expression of N-cadherin, Vimentin, and Snail in GBM cells after transfection. (J) The effect of shcircRYK and antagomir-330-5p transfection, either alone or in combination, on the formation of brain tumours was assessed using bioluminescence imaging. (K) The stemness of GSCs after transfection was measured by assessing the proportion of wells exhibiting positive results in a clonogenicity assay. (L) The expression of Oct4, Sox2, and Nanog in U87-GSC and pGBM-1-GSC cells was analysed via Western blotting after transfection; β-actin served as the loading control. Scale bar, 100 μm. Each experiment was performed thrice, and the results are displayed as the mean ± SD (**P < 0.01, ****P < 0.0001)