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MicroRNAs (miRs) are small non-coding RNAs that post-transcriptionally regulate gene expression. Recent studies indicate that miRs are important regulators of development, stem cell division, and cancer. To address the hypothesis that miR deregulation in adult neural stem cells (NSCs) is an important event in the development of human brain tumors we investigated miR expression and function in adult human gliomas and stem cells. Quantitative RT-PCR analyses of 192 microRNAs in non-tumor brain tissue (glioses), anaplastic astrocytomas, and glioblastoma multiforme (GBM) revealed six miRs, miR-7, -124a, -129, -137, -139 and -218 (hereafter glioma-miRs), that were significantly down-regulated (p<0.01; adjusted for multiple comparisons) in both primary tumor classes compared to glioses. While none of the glioma-miRs map to regions of well-established deletion in astrocytomas, miRs-124a, -129, and -137 are closely associated with CpG islands, which may act to epigenetically silence transcription. To test this, we measured glioma-miR expression in GBM cell lines (U87 and U251) treated with 5-aza-2’-deoxycytidine (5-aza-dC), a DNA methylation inhibitor, and trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor. miR-137 expression was increased up to 12-fold in cells treated with 5-aza-dC but was unchanged in cells treated with TSA. Conversely miR-129 expression was increased up to 27-fold in cells treated with TSA but not 5-aza-dC. Expression of the remaining glioma-miRs was not appreciably changed.

We also measured glioma-miR expression in cultured adult mouse neural stem cells (NSCs) induced to differentiate by mitogenic withdrawal. Expression of miR-124a and miR-137 respectively increased 8- and 24-fold by day-5, expression of miR-129 and miR-139 respectively decreased 2- and 4-fold, whereas expression of miR-7 and miR-218 did not change appreciably. Enforced overexpression of either miR-124a or miR-137 in proliferating NSCs resulted in a 5-fold increase in cells stained with the neuronal marker Tuj1 relative to controls. However the cells had distinct morphologies; miR-124a induced neuritic branching, whereas miR-137 induced a rounded appearance with no neuritic outgrowth. Co-transfection of miR-124a and -137 resulted in a ~10-fold increase in Tuj1 positive cells. Thus our results suggest that loss of miR-124a and -137 in adult NSCs impairs normal neuronal differentiation, thereby generating a pool of glial committed progenitor cells that serve as the pre-malignant seeds of gliomagenesis.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA