Definition of miRNAs expression profile in glioblastoma samples: the relevance of non-neoplastic brain reference.

PLoS One. 2013;8(1):e55314.

Visani M, de Biase D, Marucci G, Taccioli C, Baruzzi A, Pession A; PERNO Study Group.

Department of Experimental Pathology, University of Bologna, Bologna, Italy.


*Address for correspondence:

Dario de Biase, PhD BSc (

Università di Bologna

Ospedale Bellaria

Via Altura 3

40139 Bologna, Italy

Tel. N.: +390516225752 – Fax number: +390516225759



Glioblastoma is the most aggressive brain tumor that may occur in adults. Regardless of the huge improvements in surgery and molecular therapy, the outcome of neoplasia remains poor. MicroRNAs are small molecules involved in several cellular processes, and their expression is altered in the vast majority of tumors. Several studies reported the expression of different miRNAs in glioblastoma, but one of the most critical point in understanding glioblastoma miRNAs profile is the comparison of these studies. In this paper, we focused our attention on the non-neoplastic references used for determining miRNAs expression. The aim of this study was to investigate if using three different non-neoplastic brain references (normal adjacent the tumor, commercial total RNA, and epileptic specimens) could provide discrepant results. The analysis of 19 miRNAs was performed using Real-Time PCR, starting from the set of samples described above and the expression values compared. Moreover, the three different normal RNAs were used to determine the miRNAs profile in 30 glioblastomas. The data showed that different non-neoplastic controls could lead to different results and emphasize the importance of comparing miRNAs profiles obtained using the same experimental condition.

PMID: 23383149



MicroRNAs expression plays a key role in cancer development and progression and could be a target for molecular therapy. For this reason, identifying a miRNAs profile in glioblastoma (GBM) could be help in developing new therapeutic approaches. The starting material and samples used as reference control are crucial points for expression study design. Due to the difficulty of gathering non-neoplastic brain specimens there are different samples chosen as reference control in miRNAs expression analysis as normal adjacent tissues or epileptic samples. Moreover, several commercial pools of RNAs obtained from normal brain tissues were available. The present study shows that comparing miRNAs profiles obtained using different non-neoplastic controls is critical for several reasons: 1) the physiological differences in age that could be observed between different analyzed groups (e.g. usually normal adjacent the tumor samples have a mean age higher than epileptic specimens); 2) technical issues: a commercial reference is usually obtained pooling together several non-neoplastic RNAs (technical variability), while RNAs obtained from normal adjacent the tumour or epileptic specimens are not usually pooled together (biological variability); 3) different selected non-neoplastic groups could have real different miRNAs expression values. In the present study the miRNAs profiles were investigated in GBMs using the three different references (the normal area adjacent the tumor, a commercial reference, and the tissue removed in epileptic patients) as control, demonstrating that the results of miRNAs profile in brain neoplasia are strictly dependent on the non-neoplastic reference.

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