ANGI-13. TENASCIN-C INDUCES VASCULOGENIC MIMICRY FORMATION IN GLIOBLASTOMA THROUGH AKT PATHWAY

Glioblastoma is a highly invasive and vascularized primary CNS tumor. Although anti-angiogenesis therapy was effective in some tumors but not in GBM. Vasculogenic mimicry (VM), a vessel-like structure formed by highly invasive tumor cells, has been considered responsible for the failure of GBM patients for anti-angiogenesis therapy. Tenascin-c (TNC), an extracellular protein involves in tumor angiogenesis, is overexpressed in GBM. However, how TNC contributes to VM formation remains unclear. We first knocked down the expression of TNC in two glioma cell lines (U251 and A172), which had TNC overexpression. Then the formations of VM were observed by three-dimensional culture. Transwell and wound-healing assay were applied to investigate the role of TNC in cell invasion and migration abilities. The expression of MMP2, MMP9 and the phosphorylation status of AKT were determined by western-blot, RT-qPCR and gelatin zymography. Furthermore, AKT inhibitor, MK-2206 was used to block the AKT/MMP2/MMP9 signaling pathway to investigate its role in TNC regulation at different concentration (0µm, 5µm, 10µm and 20µm) for 24 hours. The knockdown efficiency was detected by western blot and RT-qPCR in U251 and A172 after siRNA transfection for 48 hours. Three-dimensional culture showed that VM-like structure formation decreased in knockdown groups compared to controls dramatically (19.67 ± 6.65 and 46.34 ± 9.10 vs. 125.67 ± 8.34 p<0.01 in U251; 59.67 ± 18.52 and 51 ± 14.97 vs. 100.67 ± 2.87, p<0.01 in A172). The invasion and migration ability of U251 and A172 were also attenuated (p<0.01) by TNC knockdown. TNC knockdown impaired the phosphorylation of AKT at both Ser473 and Thr308, and the expression of MMP2 and MMP9 were downregulated too. Exposure of AKT inhibitor blocked the VM formation as well as the expression of MMP2 and MMP9. Our study demonstrated that TNC upregulated the VM formation via AKT/MMP2/MMP9 pathway in glioma and provided a potential therapeutic target for glioblastoma by anti-angiogenesis therapy.