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MicroRNA cloning and sequencing in osteosarcoma cell lines: differential role of miR-93

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Abstract

Background

Studies show that abnormalities in non-coding genes can contribute to carcinogenesis; microRNA levels may modulate cancer growth and metastatic diffusion.

Method

MicroRNA libraries were built and sequenced from two osteosarcoma cell lines (MG-63 and 143B), which differ in proliferation and transmigration. By cloning and transfection, miR-93, expressed in both cell lines, was then investigated for its involvement in osteosarcoma progression.

Results

Six of the 19 miRNA identified were expressed in both cell lines with higher expression levels of miR-93 in 143B and in primary osteosarcoma cultures compared to normal osteoblasts. Interestingly, levels of miR-93 were significantly higher in metastases from osteosarcoma than in paired primary tumours. When 143B and MG-63 were transfected with miR-93, clones appeared to respond differently to microRNA overexpression. Ectopic expression of miR-93 more significantly increased cell proliferation and invasivity in 143B than in MG-63 clones. Furthermore, increased mRNA and protein levels of E2F1, one of the potential miR-93 targets, were seen in osteosarcoma cellular clones and its involvement in 143B cell proliferation was confirmed by E2F1 silencing.

Conclusion

Although further studies are needed to evaluate miRNA involvement in osteosarcoma progression, miR-93 overexpression seems to play an important role in osteosarcoma cell growth and invasion.

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Acknowledgements

The authors thank Paola Spessotto for transwell and invasion assays, Arcispedale Santa Maria Nuova for providing umbilical cord, Prof. Paolo Malatesta for technical support, Cristina Ghinelli for the graphic work and Alba Balladelli, B.A. for text revision. This work was supported by grants from the Italian Ministry of Health (MSB, RP), Italian Istituto Superiore della Sanità (MSB, RP), EuroBoNeT Consortium (MSB) and Institutional funds from the University of Parma (RP).

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Authors and Affiliations

  1. Center for Molecular and Translational Oncology, Department of Genetics, Microbiology, Antropology and Evolution, University of Parma, Via G. P Usberti, 11/A, 43124, Parma, Italy

    Luisa Montanini, Lisa Lasagna, Valeria Barili, Alba Murgia & Roberto Perris

  2. Department of Paediatrics, University of Parma, Via Gramsci, 14, 43124, Parma, Italy

    Luisa Montanini

  3. Department of Molecular Biology, University of Aarhus, C.F. Møllers Allé, Aarhus C, Denmark

    Søren Peter Jonstrup & Jørgen Kjems

  4. Laboratory of Experimental Oncology, Rizzoli Orthopaedic Institute, Via di Barbiano 1/10, 40136, Bologna, Italy

    Laura Pazzaglia, Amalia Conti, Chiara Novello & Maria Serena Benassi

  5. Unit for Stem Cells and Cellular Therapy, Division for Experimental Oncology 2, The National Cancer Institute Aviano, CRO-IRCCS, Via Franco Gallini 2, 33081, Aviano, PN, Italy

    Roberto Perris

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  1. Luisa Montanini
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  2. Lisa Lasagna
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Correspondence to Maria Serena Benassi.

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Montanini, L., Lasagna, L., Barili, V. et al. MicroRNA cloning and sequencing in osteosarcoma cell lines: differential role of miR-93. Cell Oncol. 35, 29–41 (2012). https://doi.org/10.1007/s13402-011-0059-z

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