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Figure 5 | Radiation Oncology

Figure 5

From: Release of monocyte migration signals by breast cancer cell lines after ablative and fractionated γ-irradiation

Figure 5

A blative γ-irradiation induces the upregulation of CD39 surface expression in MCF7 breast cancer cells. (A) CD39 surface expression on d0 and d4. Breast cancer cells were irradiated as indicated, collected by trypsinization, and CD39 surface expression was analyzed on d0 and d4 after irradiation by flow cytometry. Representative histograms are shown (black lines represent CD39 staining, filled grey areas the corresponding isotype controls). (B) Time course of CD39 upregulation. Cells were irradiated as indicated and CD39 surface expression was analyzed on d0-d4 after irradiation. Relative CD39 surface expression was calculated as the median fluorescence intensities of anti-CD39 staining subtracted by the corresponding isotype controls. Means ± s.d. of triplicates are shown. (C) Pharmacological inhibition of CD39 ectonucleotidase results in the release of monocyte migration stimulating factors by ablatively irradiated MCF7 cells. MCF7 cells were irradiated at 20 Gy or left untreated as in Figure 3A. Then, the CD39 inhibitor ARL-67156 was added at a final concentration of 100 μM and refreshed daily. The collected culture supernatants were applied to a transwell migration assay with THP-1 cells. Means ± s.d. of quadruplicates are given. (D) In silico analysis of the human CD39 promoter. Binding sites for nuclear hormone receptors (ER, PR), Egr-1, and others, including Sp-1, Stat-3 and members of the forkhead transcription factor family (Fox), were identified. (E) Analysis of p21WAF1 and Egr-1 mRNA expression in response to different irradiation regimes. Cells were irradiated as in (B), and 0–4 days after irradiation p21WAF1 and Egr-1 mRNA levels were determined by qRT-PCR analysis. Results were normalized on the means of 18S rRNA and β2-microglobulin, and untreated cells (d0) served as calibrator. Means of duplicates are given.

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