No abstract

Invasion and dissemination of medulloblastoma (MB) within the central nervous system is the principal factor predicting MB treatment failure and death. Netrin-1 is an axon guidance factor implicated in tumor and vascular biology, including in invasive behaviors. We found that exogenous netrin-1 stimulated invasion of human MB cells and endothelial cells (EC) in contrast to VEGF-A, which promoted invasion of EC but not MB cells. Further, MB cells expressed endogenous netrin-1 along with its receptors, neogenin and UNC5B. Blockades in endogenous netrin-1, neogenin or UNC5B reduced MB invasiveness. Neogenin blockade inhibited netrin-1-induced EC tube formation and recruitment of EC into Matrigel plugs, two hallmarks of angiogenesis. In pediatric MB patients, netrin-1 mRNA levels were increased 1.7-fold in MB tumor specimens compared to control specimens from the same patient. Immunohistochemical analyses showed that netrin-1 was elevated in MB tumors versus cerebellum controls. Notably, urinary levels of netrin-1 were 9-fold higher in MB patients compared to control individuals. Moreover, urinary netrin-1 levels were higher in patients with invasive MB compared to patients with non-invasive MB. Lastly, we noted that urinary netrin-1 levels diminished after MB resection in patients. Our results suggest netrin-1 as a candidate biomarker capable of detecting an invasive, disseminated phenotype in MB patients and predicting their disease status.

The purpose of this study was to investigate the functions of microRNA-9, which is a tissue-specific microRNA in central nervous system, in the vasculogenic mimicry (VM) of glioma cell lines in vitro and in vivo. Glioma cell lines U87MG, U251 and SHG44 were transfected with microRNA-9 mimic, microRNA-9 inhibitor or scramble sequences. The amount of microRNA-9 and Stathmin (STMN1) mRNA was determined by quantitative real-time PCR, and the protein expression of STMN1 was determined by western blot. Cell proliferation and apoptosis were assessed. The interactions between the 3'UTR of STMN1 and miR-9 was determined by luciferase reporter assay. The VM capacity in vitro was evaluated using VM formation assay, and the rescue experiment of STMN1 was carried out in U251 cells. The in vivo experiment was applied with animal models implanted with U87MG cells.MicroRNA-9 mimic transfection reduced proliferation and increased apoptosis in glioma cell lines (p < 0.05). MicroRNA-9 mimic up-regulated STMN1 mRNA levels but reduced its protein levels (p < 0.05), and luciferase activity of STMN1 was suppressed by microRNA-9 mimic transfection (p < 0.05). Furthermore, microRNA-9 mimic transfection suppressed tumor volume growth, as well as VM both in vitro and in vivo. The cell viability and microtube density were upregulated in U251 cells after STMN1 up-regulation (p < 0.05). STMN1 is a target of microRNA-9, and microRNA-9 could modulate cell proliferation, VM and tumor volume growth through controlling STMN1 expression. MicroRNA-9 and its targets may represent a novel panel of molecules for the development of glioma treatment.