- - - Department of Medical Physics and Biomedical Engineering, Tehran University of Medical Sciences, Tehran, Iran
- - - Department of Medical Genetics, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
- - - Department of Medical Physics and Biomedical Engineering, Tehran University of Medical Sciences, Tehran, Iran
- - - Department of Bioscience and Biotechnology, Malek-Ashtar University of Technology, Tehran, Iran
- - - Department of Medical Genetics, Medical Biotechnology Institute, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran
- - - Radiation Oncology Research Center, Cancer Institute, Tehran University of Medical Sciences, Tehran, Iran

Background: The paradigm shifts in target theory could be defined as the radiation-triggered bystander response in which the radiation deleterious effects occurred in the adjacent cells. Objective: This study aims to assess bystander response in terms of DNA damage and their possible cell death consequences following high-dose radiotherapy. Temporal characteristics of gH۲AX foci as a manifestation of DNA damage were also evaluated.Material and Methods: In this experimental study, bystander response was investigated in human carcinoma cells of HeLa and HN۵, neighboring those that received high doses. Medium transfer was performed from ۱۰ Gy-irradiated donors to ۱.۵ Gy-irradiated recipients. GammaH۲AX foci, clonogenic and apoptosis assays were investigated. The gH۲AX foci time-point study was implemented ۱, ۴, and ۲۴ h after the medium exchange. Results: DNA damage was enhanced in HeLa and HN۵ bystander cells with the ratio of ۱.۲۷ and ۱.۷۲, respectively, which terminated in more than two-fold clonogenic survival decrease, along with gradual apoptosis increase. GammH۲AX foci temporal characterization revealed maximum foci scoring at the ۱ h time-point in HeLa, and also ۴ h in HN۵, which remained even ۲۴ h after the medium sharing in higher level than the control group.  Conclusion: The time-dependent nature of bystander-induced gH۲AX foci as a DNA damage surrogate marker was highlighted with the persistent foci at ۲۴ h. considering an outcome of bystander-induced DNA damage, predominant role of clonogenic cell death was also elicited compared to apoptosis. Moreover, the role of high-dose bystander response observed in the current work clarified bystander potential implications in radiotherapy.

Bystander, High-Dose, GammaH۲AX, DNA Damage, Survival, apoptosis