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BMC Cancer
Published in: BMC Cancer 1/2014

Open Access 01-12-2014 | Research article

Deep Sequencing the microRNA profile in rhabdomyosarcoma reveals down-regulation of miR-378 family members

Authors: Francesca Megiorni, Samantha Cialfi, Heather P McDowell, Armando Felsani, Simona Camero, Alessandro Guffanti, Barry Pizer, Anna Clerico, Alessandra De Grazia, Antonio Pizzuti, Anna Moles, Carlo Dominici

Published in: BMC Cancer | Issue 1/2014

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Abstract

Background

Rhabdomyosarcoma (RMS) is a highly malignant tumour accounting for nearly half of soft tissue sarcomas in children. MicroRNAs (miRNAs) represent a class of short, non-coding, regulatory RNAs which play a critical role in different cellular processes. Altered miRNA levels have been reported in human cancers, including RMS.

Methods

Using deep sequencing technology, a total of 685 miRNAs were investigated in a group of alveolar RMSs (ARMSs), embryonal RMSs (ERMSs) as well as in normal skeletal muscle (NSM). Q-PCR, MTT, cytofluorimetry, migration assay, western blot and immunofluorescence experiments were carried out to determine the role of miR-378a-3p in cancer cell growth, apoptosis, migration and differentiation. Bioinformatics pipelines were used for miRNA target prediction and clustering analysis.

Results

Ninety-seven miRNAs were significantly deregulated in ARMS and ERMS when compared to NSM. MiR-378 family members were dramatically decreased in RMS tumour tissue and cell lines. Interestingly, members of the miR-378 family presented as a possible target the insulin-like growth factor receptor 1 (IGF1R), a key signalling molecule in RMS. MiR-378a-3p over-expression in an RMS-derived cell line suppressed IGF1R expression and affected phosphorylated-Akt protein levels. Ectopic expression of miR-378a-3p caused significant changes in apoptosis, cell migration, cytoskeleton organization as well as a modulation of the muscular markers MyoD1, MyoR, desmin and MyHC. In addition, DNA demethylation by 5-aza-2′-deoxycytidine (5-aza-dC) was able to up-regulate miR-378a-3p levels with a concomitant induction of apoptosis, decrease in cell viability and cell cycle arrest in G2-phase. Cells treated with 5-aza-dC clearly changed their morphology and expressed moderate levels of MyHC.

Conclusions

MiR-378a-3p may function as a tumour suppressor in RMS and the restoration of its expression would be of therapeutic benefit in RMS. Furthermore, the role of epigenetic modifications in RMS deserves further investigations.
Appendix
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Metadata
Title
Deep Sequencing the microRNA profile in rhabdomyosarcoma reveals down-regulation of miR-378 family members
Authors
Francesca Megiorni
Samantha Cialfi
Heather P McDowell
Armando Felsani
Simona Camero
Alessandro Guffanti
Barry Pizer
Anna Clerico
Alessandra De Grazia
Antonio Pizzuti
Anna Moles
Carlo Dominici
Publication date
01-12-2014
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2014
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/1471-2407-14-880

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