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. 2011 Sep 12;6(9):e24584. doi: 10.1371/journal.pone.0024584

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de Antonellis et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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A. Differentiating effects of AdV-GFP-miR-34a on tumor spheres from Patch^+/- P53^+/- mice. Representative microscopy images and confocal GFP immunofluorescence staining of Patch^+/- P53^+/- mouse MB spheres at 24 h and 96 h from AdV-GFP-mock (left) or AdV-GFP-miR-34a (right) virus infections. B. Left: Real-time PCR analysis showing expression levels of the neural markers Nestin, MAP2, TUJ1, and GFAP in Patch^+/- P53^+/- tumor spheres at 96 h from infection with AdV-miR-34a or AdV-mock viruses. Fold changes are shown, calculated with respect to the gene expression of the AdV-mock infected tumor-spheres. Data are means ±standard deviations of 3 experiments, each carried out in triplicate. Real-time PCR reactions were normalized to β-Actin. Right: Representative immunofluorescence staining of Patch^+/- P53^+/- tumor spheres, differentiated following viral delivery of miR-34a, performed using an anti-GFAP antibody. GFP indicates the infection efficiency and the tumor sphere viability. C. Doxorubicin treatment of Patch^+/- P53^+/- and Patch^+/- P53^-/- tumor spheres. Representative microscopy images showing the neural differentiating phenotype observed only for the P53^+/- tumor spheres. D. Left: Representative confocal GFP immunofluorescence staining of Patch^+/- P53^+/- tumor spheres at 24 h from AdV miR-34a virus infection. Arroweds denotes that the AdV-miR-34a virus efficiently infects only the cells located in the most external regions of the tumor spheres. Right: Illustration of the cell z-slices imaged.