Elsevier

Brain Research

Volume 1236, 21 October 2008, Pages 185-193
Brain Research

Research Report
hsa-mir-181a and hsa-mir-181b function as tumor suppressors in human glioma cells

https://doi.org/10.1016/j.brainres.2008.07.085Get rights and content

Abstract

MicroRNAs (miRNAs) are small noncoding RNA molecules that regulate protein expression by cleaving or repressing the translation of target mRNAs. In mammal animals, their function mainly represses the target mRNAs transcripts via imperfectly complementary to the 3′ UTR of target mRNAs. Several miRNAs have been recently reported to be involved in modulation of glioma development, especially some up-regulated miRNA, such as hsa-miR-21 and hsa-miR-221. However, here we reported that the down-regulated hsa-miR-181a and hsa-miR-181b of hsa-miR-181 family were also involved in oncogenesis of glioma. Our studies showed that hsa-miR-181a and hsa-miR-181b functioned as tumor suppressors which triggered growth inhibition, induced apoptosis and inhibited invasion in glioma cells. Furthermore, the tumor-suppressive effect of hsa-miR-181b in glioma cells was more apparent than the effect of hsa-miR-181a. These findings suggest aberrantly down-regulated hsa-miR-181a and hsa-miR-181b may be critical factors that contribute to malignant appearance in human gliomas.

Introduction

MicroRNAs (miRNAs) are located in noncoding regions or the introns of the genome. Mature functional miRNAs of approximately 22 nucleotides generated from long primary miRNA (pri-miRNA) transcripts control gene expression at the post-transcriptional level by degrading or repressing target mRNAs. miRNAs are evolutionarily selected gene regulatory molecules. Each type of cell is likely to have a specific miRNA milieu to control gene expression (Chen, 2005). And each miRNA has the potential to regulate about 200 target genes according to recent computational predictions. It implies that over one third of protein-coding genes in humans are regulated by miRNAs. Thus, miRNA-mediated gene regulation is now considered an important role in biologic processes. They are found to regulate apoptosis, proliferation, differentiation, development, and metabolism in worm, fly, fish, mouse, and human cells (Bartel, 2004).

Recently, some miRNAs aberrantly expressed in cancer have been well documented. They were found to regulate the expression of signaling molecules, such as cytokines, growth factors, transcription factors, and proapoptotic and antiapoptotic genes. And some miRNAs may function as tumor suppressors or oncogenes involved in the pathogenesis of tumors by targeting oncogenes or tumor-suppressor genes (Chen, 2005). In this regard, tumor-suppressive miRNAs are usually underexpressed in tumors, and the reduction in the expression may cause increased expression of the oncogenic target genes. For instance, let-7 miRNA acts as tumor-suppressor genes, down-regulated in lung tumors and associated with a poor postoperative prognosis (Junichi et al., 2004). The RAS oncogene is proved to be regulated by the let-7 miRNA; decreased expression of let-7 miRNA in lung tumors causes increased expression of the RAS oncogene and thereby may contribute to tumorigenesis (Johnson et al., 2005). There is also evidence that miR-17-92 cluster acts as an oncogene in many malignant lymphomas containing amplified 13q31–32 chromosomal fragments (He et al., 2005). And miR-17-92 cluster can augment the oncogenic effect of the c-Myc gene in mice (Lu et al., 2007). Thus, oncogenes and tumor-suppressor genes could be potentially regulated by miRNAs, and miRNAs are presumed to be a class of genes involved in human tumorigenesis.

In human glioblastoma cells, Chan et al reported miR-21 over-expressed in glioblastoma cells, and acted as an antiapoptotic factor (Chan et al., 2005). In this study, we analyzed the function of hsa-miR-181a and hsa-miR-181b in human gliomas and glioma cell lines, which are enriched in normal brain but sharply down-regulated in human gliomas. Our data showed hsa-miR-181a and hsa-miR-181b function as tumor-suppressor factors in human glioma.

Section snippets

hsa-miR-181a and hsa-miR-181b were down-regulated in human gliomas cells

To investigate whether hsa-miR-181a and hsa-miR-181b were down-regulated in human gliomas and glioma cell lines as Ciafre et al. reported in ref. Ciafre et al. (2005), we performed the TaqMan-based real-time stem–loop RT-PCR analyses. Our data showed similar results that hsa-miR-181a and hsa-miR-181b were strongly down-regulated in all grade glioma samples (WHO-II, WHO-III and WHO-IV glioma tissues) versus normal brain tissues, and hsa-miR-181a expression was negatively correlated with tumor

Discussion

It is now well known that hsa-miR-181 family, one of the best-studied miRNA families, dysregulated in many tumors. However, extremely little information has been reported to their physiological and pathological roles in tumors. In this regard, we have reported hsa-miR-181a and hsa-miR-181b were down-regulated in human gliomas and glioma cell lines. Moreover, we have firstly identified hsa-miR-181a expression negatively correlates with tumor grade in gliomas. Furthermore, over-express

Cell lines and culturing conditions

Human glioma cell lines, U87, TJ905, and U251, were purchased from Chinese Academy of Sciences Cell Bank. All glioma cell lines were maintained in a 37 °C, 5% CO2 incubator in DMEM supplemented with 10% fetal bovine serum (FBS), and routinely passaged at 2- to 3-day intervals. And experiments were divided into four groups as blank control group, miRNA scrambled group, has-miR-181a group and has-miR-181b group.

RNA isolation

Human glioma tissue samples were obtained from the first affiliated hospital of

Acknowledgments

We thank Drs X.H. Ji and C.S. Kang for experimental support, discussions and suggestions. We are grateful to Genesil Company for miRNA reporter vectors constructs. This work is supported by the China Natural Science Foundation (Proj. No. 30672165 and 30772231) and Jiangsu Province's Medical Major Talent program (Proj. No. RC2007061).

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    Contributed equally.

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