Single Shot Irradiation and Molecular Effects on a Diffuse Large B Cell Lymphoma Cell Line

Fernando Mendes^{1, 2,} , Cátia Domingues^{3, 4}, Susann Schugk⁵, Ana Margarida Abrantes^{1, 3}, Ana Cristina Gonçalves^{3, 4}, Tiago Sales¹, Ricardo Teixo¹, Rita Silva¹, Jéssica Estrela⁶, Mafalda Laranjo^{1, 3}, João Casalta-Lopes^{1, 7}, Clara Rocha^{8, 9}, Paulo César Simões⁷, Ana Bela Sarmento^{3, 10}, Maria Filomena Botelho^{1, 3} and Manuel Santos Rosa¹¹

¹Biophysics and Biomathematics Institute, IBILI-Faculty of Medicine, University of Coimbra, Portugal

²Polytechnic Institute of Coimbra, ESTESC-Coimbra Health School, Department Biomedical Laboratory Sciences, Coimbra, Portugal;CIMAGO, FMUC-Faculty of Medicine, University of Coimbra, Portugal;CNC.IBILI, Universidade de Coimbra, Portugal

³CIMAGO, FMUC-Faculty of Medicine, University of Coimbra, Portugal

⁴Oncobiology and Hematology Laboratory Applied Molecular Biology and Clinical University of Hematology, Faculty of Medicine of University of Coimbra, Coimbra, Portugal

⁵Biomedical Laboratory Sciences Department, University of Gothenburg

⁶Polytechnic Institute of Coimbra, ESTESC-Coimbra Health School, Department Biomedical Laboratory Sciences, Coimbra, Portugal

⁷Radiation Oncology Department, Hospital and University Center of Coimbra, Coimbra, Portugal

⁸Polytechnic Institute of Coimbra, ESTESC-Coimbra Health School, Department Complementary Sciences, Coimbra, Coimbra, Portugal

⁹Institute for Systems Engineering and Computers at Coimbra, Coimbra, Portugal

¹⁰Oncobiology and Hematology Laboratory Applied Molecular Biology and Clinical University of Hematology, Faculty of Medicine of University of Coimbra, Coimbra, Portugal;Hematology Clinic Department, Centro Hospitalar Universitário de Coimbra, Coimbra, Portugal

¹¹Immunology Institute, Faculty of Medicine, University of Coimbra, Coimbra, Portugal

Cite this paper:
Fernando Mendes, Cátia Domingues, Susann Schugk, Ana Margarida Abrantes, Ana Cristina Gonçalves, Tiago Sales, Ricardo Teixo, Rita Silva, Jéssica Estrela, Mafalda Laranjo, João Casalta-Lopes, Clara Rocha, Paulo César Simões, Ana Bela Sarmento, Maria Filomena Botelho and Manuel Santos Rosa. Single Shot Irradiation and Molecular Effects on a Diffuse Large B Cell Lymphoma Cell Line. Journal of Cancer Research and Treatment. 2016; 4(1):9-16. doi: 10.12691/jcrt-4-1-2

Abstract

Background: Diffuse large B cell lymphoma (DLBCL) is recognized as a heterogeneous group of hematological malignancies, and collectively forms the most common type of aggressive, non-Hodgkin lymphoma. The aim of our study was to evaluate the cellular and molecular effects of high doses X radiation in a DLBCL cell line. Materials and Methods: Farage cells were cultured and exposed to 0.5-60 Gy of Ionizing Radiation (IR). Cell viability and proliferation were assessed by trypan blue assay. Cell survival was determined with clonogenic assay. Cell death was assessed by flow cytometry (FC) and by optical microscopy. Cell cycle, mitochondrial membrane potential, reactive oxygen species, GSH and BAX/BCL-2 ratio were measured by FC. DNA damage was evaluated using comet assay. Total and phosphorylated P53 was assessed by western blot. Results: IR induced cytotoxic and cytostatic effects in Farage cells in a dose and time dependent manner with an LD50 of 1.73 Gy. Cell death occurs mainly by apoptosis or later apoptosis/necrosis with an increase in BAX/BCL-2 ratio and a significant increase in ROS production. We also observed cell cycle arrest at G₂/M phase and a significant increase in DNA damage as well as P53 total and phosphorylated expression levels. Conclusions: High doses of IR induces a time and dose dependent response which leads to increased ROS production, DNA damage with the increased P53 expression and activation expressed by elevated levels of pP53, resulting in G₂/M cell cycle arrest and increases in later apoptosis/necrosis cell death. Our results showed that single shot IR induces effects in different cell components and its comprehension is essential to choose the treatment planning.

Keywords:
large diffuse B cell lymphoma radiotherapy genotoxicity oxidative stress cell death P53

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