Radiobiological Response Of Breast Cancer Cells, Exposed To Atmospheric Pressure Plasmas And Ionizing Radiation, An In-Vitro Essay. (Project Overview)

The purpose of this project is to describe the application of an ionized gas to different lines of breast cancer cells, added to application of a clinical photon beam, by measuring variations in cell death rate. This procedure may lead to implementation of a new in vitro technique for studying new radiobiological approaches for adjuvant methods to radiotherapy techniques.

Characterization of the plasma generator

An in-house developed Cold Atmospheric Plasma "CAP" will be evaluated in terms of its functionality and features in geometry and operation. The characteristics of the plasma generated using an Argon-Helium gas mixture, should be analyzed regarding its possibilities of interaction with a biological medium, depending on properties such as the Power of the source; Gas pressure; Visible and infrared emission wavelength by FTIR Microscopy and Ultra-Violet (UVA, UVB, UVC) by UV absorption spectroscopy [1]; Gas temperature; Exposure time; Determination of reactive species, and Gas flow (5 l/min).

Biological Evaluations

The cell lines of human breast adenocarcinoma (MCF-7, MDA-MB-231, MDA-MB-468, Hs 578T, T-47D, MDA-N and BT-54), will be placed in plates of deposits with regular flat bottom for the application of ionizing radiation and CAP, using a cell culture medium; RPMI 1640 supplemented with 10% fetal bovine serum, 100 units/ml penicillin, 100 μg/ml streptomycin and 0.25 μg/ml amphotericin B, kept at 37 ° C, in an atmosphere humidified with 5% CO₂ and 95% air. In order to assess the cell viability at different radiation doses, in the presence and absence of treatment with CAP, the samples will be analyzed with the DNA binding fluorochromes Hoechst 33342 (slightly compromised cell membrane), and Propidium Iodide (complete compromised cell membrane). Images will be acquired with the multi-modal Cytation™3 microplate reader and analyzed for segmentation and quantification of the cell nuclei for each of the fluorescent components, with Cell Profiler software.

Dosimetric measurements

For dose delivery, a beam of 6 MV from a Varian CLINAC iX (Varian Medical Systems, Palo Alto, California) will be used. This beam was calibrated in water following the protocol TECDOC-398. For the determination of dose levels, radio-chromic films (4x4 cm² samples) will be irradiated and measured for calibration using a spectrophotometer in the reflection mode [2].

Expected results

Cellular modifications produced by the combining of cold plasma characteristics and ionizing radiation, will generate a statistical sample that may allow us to determine the overall effect of modification in radiosensitivity by the exposure to the CAP [3]. In order to define the variation rate in which the plasma applied to cancer cells, enhance programmed cell death, this could increase the uniformity in response of the cell group to ionizing radiation, which may be a good contribution in validation of clinical procedures against cancer with greater effectiveness.