Establishment of animal model for the analysis of cancer cell metastasis during radiotherapy

Background: Gamma-Ionizing radiation (IR) therapy is one of major therapeutic tools in cancer treatment. Nevertheless, gamma-IR therapy failed due to occurrence of metastasis, which constitutes a significant obstacle in cancer treatment. The main aim of this investigation was to construct animal model which present metastasis during radiotherapy in a mouse system in vivo and establishes the molecular mechanisms involved. Materials and methods: The C6L transfectant cell line expressing firefly luciferase (fLuc) was treated with gamma-IR, followed by immunoblotting, zymography and invasion assay in vitro. We additionally employed the C6L transfectant cell line to construct xenografts in nude mice, which were irradiated with gamma-IR. Irradiated xenograft-containing mice were analyzed via survival curves, measurement of tumor size, and bioluminescence imaging in vivo and ex vivo. Metastatic lesions in organs of mice were further assessed using RT-PCR, H & E staining and immunohistochemistry. Results: gamma-IR treatment of C6L cells induced epithelial-mesenchymal transition (EMT) and increased cell invasion. In irradiated xenograft-containing mice, tumor sizes were decreased dramatically and survival rates extended. Almost all non-irradiated xenograft-containing control mice had died within 4 weeks. However, we also observed luminescence signals in about 22.5% of gamma-IR-treated mice. Intestines or lungs of mice displaying luminescence signals contained several lesions, which expressed the fLuc gene and presented histological features of cancer tissues as well as expression of EMT markers. Conclusions: These findings collectively indicate that occurrences of metastases during gamma-IR treatment accompanied induction of EMT markers, including increased MMP activity. Establishment of a murine metastasis model during gamma-IR treatment should aid in drug development against cancer metastasis and increase our understanding of the mechanisms underlying the metastatic process.