Phenotypic similarities have long been recognized between subpopulations of glioma cells

Phenotypic similarities have long been recognized between subpopulations of glioma cells and neural stem cells. regulator of the development of stem cell-like phenotypes and cell motility in malignant glioma cells. < 0.05 in all cases was considered statistically significant. Results miR-204 was down-regulated in gliomas To identify miRNAs with similar expression patterns in glioma and neural stem cells, we retrieved miRNA expression profiles from two datasets, The Cancer Genome Atlas (TCGA) and the GEO dataset "type":"entrez-geo","attrs":"text":"GSE29759","term_id":"29759"GSE29759. As shown in Fig. 1A, miR-204 emerged as one of the most prominent down-regulated miRNAs in the TCGA glioblastoma (GBM) dataset, accompanied by several other down-regulated miRNAs reported previously, namely, miR-124, 106a, Carebastine IC50 17C5p and members of the miR-181 family (15, 16). As documented previously, miRNAs up-regulated in gliomas, including miR-21 (17C19), miR-9 (20, 21) and miR-10b, were also screened out. Carebastine IC50 (22, 23). Comparing the miRNA profiles in the TCGA dataset with those in neural stem cells identified that miR-204 was down-regulated in both glioma and neural stem cells (Fig. 1B). To validate the expression pattern of miR-204 in gliomas, qRT-PCR was conducted with Carebastine IC50 6 normal brain tissue samples and 20 freshly dissected glioma samples. In consistence with the data presented in the TCGA profile, expression of miR-204 was markedly down-regulated in high-grade glioma samples (WHO tumor grade III and IV) and to a lesser degree, decreased in WHO tumor grade I and II glioma samples, as compared to normal brain tissues (Fig. 1C). Of note, 10 out of 14 glioma cell lines displayed significantly down-regulated expression of miR-204 relative to that in two primary cultured normal human astrocytes Carebastine IC50 (NHA) (Fig. 1D). Figure 1 miR-204 expression is down-regulated in glioma and neuronal stem cells Restoration of miR-204 in glioma cells suppressed cellular migration and invasion To investigate whether miR-204 could modulate the migration and invasiveness of glioma cells, SNB19 and LN382T glioma cell lines, which possessed the lowest levels of miR-204 expression among all tested glioma cell lines, were chosen for further studies. By retrovirally transducing glioma cells with miR-204 at expression levels comparable with that of NHAs (Fig. S1), we first examined the effect of miR-204 on cell migration using wound healing assay. As shown in Fig. 2A, compared with the vector-control cells, which spread to the center line within 16 hours, miR-204-transduced cells exhibited considerably slower migration and reduced cell spreading. Furthermore, we tracked the movement of glioma cells, using a time-lapse imaging system, and examined whether miR-204 altered the migrating behavior of individual glioma cells. As shown in Fig. 2B, miR-204 transduced-SNB19 and -LN382T cells generally retained their original positions, whereas vector-control cells moved away at higher extents. Specifically, we measured the migration tracks of 10 individual cells within 5 hours and observed that miR-204 transduced cells moved within a very limited area. In contrast, the control cells spread out in a markedly larger area, with approximately 1.5C3 folds of increase in migration distance (Fig. 2C). To further determine the effect of miR-204 expression on motility of glioma cells, transwell penetration assay were performed. As shown in Carebastine IC50 Fig. 2D, less number of miR-204 cells invaded across the membrane pre-coated with matrigels when compared with control cells. Importantly, the three-dimensional spheroid invasion assay showed that vector-control cells displayed a highly aggressively penetrating growth after 5 days, whereas the miR-204 transduced-cells grew in spherical colonies (Fig. 2E). Taken Rabbit polyclonal to TDGF1 together, these observations suggested that miR-204 strongly.

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