Investigation on the Direct and Bystander Effects in HeLa and CHO Cells Exposed to Very Low Dose Alpha-Radiation Using Electrical Impedance Measurement
Student thesis: Doctoral Thesis
Related Research Unit(s)
Radiation-induced bystander effects (RIBE) have raised many concerns about radiation safety and protection. In RIBE, unirradiated cells receive signals from irradiated cells and exhibit irradiation effects. Until now, most RIBE studies have been based on morphological and biochemical characterization. However, research on the label-free study of RIBE impact on the biophysical properties of cells has been lagging. Furthermore, the distortion of the output signal by cell mobility has been the significant challenge minimizing the amount of information acquired from the well-known label-free technique like impedance spectroscopy. Here, electrical cell indium tin oxide (ITO) substrate impedance spectroscopy (ECIIS) has been used as bioimpedance sensors to monitor cell behaviours. The ITO substrate needed for proper cell adherence was prepared by stepwise treatment and characterization of aminosilanized-ITO with N-Hydroxy succinimide (NHS) and 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide-hydrochloride (EDC) to create a carboxyl group on its surface. The impact of the ITO treatment on adherent cell mobility and suitability in ECIIS real-time analysis of cell behaviour was also investigated. Furthermore, the ECIIS system was modelled and analyzed with simple equations to understand the relationship between the system parameters and cell sensing properties. Lastly, the ECIIS was employed to study RIBE in Chinese hamster ovary (CHO) cells and human cervical carcinoma (HeLa) cells. For both cell types, the bioimpedance of bystander cells, alpha(α)-particle (Am-241) irradiated, and untreated/unirradiated cells were monitored with a sampling interval of 8 seconds over a period of 24 hours. Media from irradiated cells exposed to different radiation doses were used to investigate the radiation dose dependence of bystander cells. In parallel, images of the cells were taken at times where impedance changes were found. A surface roughness of 6.37 nm with enhanced conductivity, 75% visible light transparency, improved cell adherence and reduced collective cell migration speed by ~2 fold was observed in the carboxyl-terminated ITO substrate with diminished signal distortion when utilized in the ECIIS. By analyzing the changes in absolute impedance and cell size/cell number with time, it is observed that bystander cells mimicked irradiated cells in terms of modification in cell morphology and proliferation rate. Furthermore, these effects appeared to be time-dependent and inversely proportional to the radiation dose. Thus, this study provides an ITO surface-treatment strategy to reduce multiple cell movement effects and obtained essential cell information from the ECIIS study of multiple cells. Furthermore, this approach allows a label-free analysis of cellular responses to RIBE with high sensitivity and temporal resolution and providing crucial insights into the RIBE mechanism.
- Biophysics, Radiotherapy, Electrical impedance spectroscopy, Biofunctionalization