Regulation of IL-8 gene expression in gliomas by microRNAs

Interleukin-8 (IL-8, or CXCL8) is a major promoter of angiogenesis and invasiveness in human gliomas, where it is expressed and secreted at high levels. Among the different control levels of IL-8 gene expression in gliomagenesis, several have been studied and well characterized, such as hypoxia/anoxia stimulation, response to Fas ligation, death receptor activation, cytosolic Ca2+ transients, TNF-alpha, IL-1, and other cytokines and various cellular stresses. In addition, the expression of the IL-8 gene might be under the control of epigenetic mechanisms(s), such as those regulated by microRNAs. We found that bacterial challenge, which is known to strongly activate IL-8 gene transcription in epithelial cells, is downregulated by miR-93 (Fabbri, 2014). This is of peculiar interest for cancerogenesis since miR-93 has been found involved in the downregulation of expression of VEGF which cooperates, together with IL-8, in glioma angiogenesis. Expression levels of IL-8 and VEGF genes and of miR-93, have been investigated in High Grade and Low Grade Gliomas (HGG and LGG) samples and in U251 human glioma cells. Pre-miR- and anti-miR-93 were transfected in U251 cells to check modulation of candidate target genes (IL-8 and other cytokines relevant to the glioma microenvironment). Both VEGF and IL-8 mRNAs were higly expressed in LGGs (20-200 folds) and HGGs (20-300 folds) in respect to reference RNA from healthy brains. VEGF and IL-8 mRNAs expression correlated directly, whereas MiR-93 correlated inversely with both target genes transcripts in glioma specimens. In silico analyses evidenced consensus sequences for the interaction of miR-93 in the 3'-UTR regions of IL-8 and VEGF genes. Transfection of U251 glioma cells with pre-miR-93 down-modulated both VEGF and IL-8 genes expression, whereas anti-miR-93 resulted in a consistent up-modulation. Our results suggest that microRNAs, including miR-93, might be proposed as relevant post-transcriptional regulators of angiogenesis in human gliomas.