Enzymatic Spermine Metabolites Induce Apoptosis Associated with Increase of p53, caspase-3 and miR-34a in Both Neuroblastoma Cells, SJNKP and the N-Myc-Amplified Form IMR5

酶促精胺代谢物诱导神经母细胞瘤细胞 SJNKP 和 N-Myc 扩增形式 IMR5 中的细胞凋亡,并与 p53、caspase-3 和 miR-34a 的增加有关

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作者:Yuta Kanamori, Alessia Finotti, Laura Di Magno, Gianluca Canettieri, Tomoaki Tahara, Fabio Timeus, Antonio Greco, Paola Tirassa, Jessica Gasparello, Pasquale Fino, Carlo Maria Di Liegro, Patrizia Proia, Gabriella Schiera, Italia Di Liegro, Roberto Gambari, Enzo Agostinelli

Abstract

Neuroblastoma (NB) is a common malignant solid tumor in children and accounts for 15% of childhood cancer mortality. Amplification of the N-Myc oncogene is a well-established poor prognostic marker in NB patients and strongly correlates with higher tumor aggression and resistance to treatment. New therapies for patients with N-Myc-amplified NB need to be developed. After treating NB cells with BSAO/SPM, the detection of apoptosis was determined after annexin V-FITC labeling and DNA staining with propidium iodide. The mitochondrial membrane potential activity was checked, labeling cells with the probe JC-1 dye. We analyzed, by real-time RT-PCR, the transcript of genes involved in the apoptotic process, to determine possible down- or upregulation of mRNAs after the treatment on SJNKP and the N-Myc-amplified IMR5 cell lines with BSAO/SPM. The experiments were carried out considering the proapoptotic genes Tp53 and caspase-3. After treatment with BSAO/SPM, both cell lines displayed increased mRNA levels for all these proapoptotic genes. Western blotting analysis with PARP and caspase-3 antibody support that BSAO/SPM treatment induces high levels of apoptosis in cells. The major conclusion is that BSAO/SPM treatment leads to antiproliferative and cytotoxic activity of both NB cell lines, associated with activation of apoptosis.

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