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Subramanian Krishnakumar, Nalini Venkatesan, Vikas Khetan, Maddy Reddy; Role of miR 106 - 25 family in Retinoblastoma tumouriogenesis: in-vitro analysis of their functions using antagomirs. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3969.
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Retinoblastoma (RB), a primary pediatric intraocular tumor. Several novel molecular strategies are being developed for the clinical management of RB. Here, the role of selected miRNA cluster: miR106b~25 family have been analyzed for potential diagnostic and therapeutic targeting in RB.
The miRNA cluster (miR-106b, miR-93, miR-25) and their direct regulatory gene , Miniature chromosome maintenance 7(MCM7) complex were confirmed by quantitative Reverse Transcriptase PCR (qRT-PCR) in primary retinoblastoma tissues (N= 21).Functional role of these miRNA cluster have been studied by using antagomirs in cultured RB (Y79,Weri Rb-1) cells in-vitro. The expression of gene targets - p21 / BIM regulated by miR-106b family, was confirmed by western blot. Cell proliferation and apoptotic studies have been performed using Cell viability assay (MTT), Colony forming assay (Anchorage dependent assay), and Annexin V binding assay, in the antagomirs-treated RB cells (Y79 and Weri Rb-1).
In the 21 tumor tissues analysed, miR-106b was detected in 90.47%, miR-93 in 85.71%, miR-25 in 95.23% respectively. In association with these miRNA clusters, we observed higher expression of MCM7 in the RB primary tumors. In-vitro silencing of these miRNAs in RB cells resulted in the reduction of cell proliferation and in colony formation, through the induction of apoptosis (confirmed by Annexin V assay).
The aberrant expression of miR-106b-25family in primary retinoblastoma implicates its role in RB tumorigenesis. Diagnostic/ Prognostic potential for miR 106b ~25 family miRNAs in RB is indicated. The reduction in cancer cell viability by specific antagomir suggests a potential target for RB therapy. Funding agency: Childhood Eye cancer trust (CHECT Foundation), London, UK
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