Zebrafish irradiation: The effects of high dose-rate radiation on zebrafish embryos

Finding a favorable cure for cancer has been one of the main clinical challenges today. Nowadays the majority of the patients is treated with radiotherapy. Recently research in to a new paradigm of radiotherapy, so called FLASH radiotherapy, has opened up a new insight in to reducing negative side effects. FLASH dose rates (>40 Gy/s) have the ability to reduce the prevalence of negative side effects in patients without sacrificing tumor control. This study focuses on whether dose rate affects the degree of damage to healthy tissues caused by ionizing radiation. Understanding the biological mechanisms that influence the response to radiotherapy and specifically FLASH radiotherapy is therefore key knowledge in the development of new therapeutic protocols.This study evaluates dose-rate effects using zebrafish embryos as a small model organism. To study dose-rate effects the zebrafish are irradiated at 24 hours post fertilization and biomarkers are researched to asses dose-rate effects. They are irradiated using a Co60 irradiator providing a dose-rate of 0.015 Gy/s and a second one providing a dose-rate of 0.211 Gy/s. Several biomarkers for radiation damage are identified, including rate of embryonic development (hatching), DNA breaks and apoptosis. The DNA breaks are measured using a TUNEL assay and apoptosis through a caspase-3 assay. The results of the assay's are measured using confocal microscopy.Monte Carlo simulations of the experimental setup were performed to assess the uniformity of radiation and effective dose. They show a homogeneous dose throughout the sample, they show a 37% lower dose in water in the sample compared to air.The hatching biomarker shows that the hatching rates are 0.24 +-0.03 for non irradiated samples and 0.37 +- 0.31 and 0.43 +- 0.20 for samples that have been irradiated with respectively 0.008 Gy/s and 0.015 Gy/s with a total dose of 10 Gy. It shows that the hatching rates decreases for the irradiated embryos compared to the non irradiated samples however no significant difference can be found for different dose-rates. The sub-cellular biomarkers, the TUNEL assay and the caspase-3 assay, seem more promising in detecting dose-rate differences but more research is needed to find the dose-rate effects on DNA damage and apoptosis. In order to gain a fuller picture into the biological mechanisms of FLASH dose-rate radiation a greater variety of biomarkers and more research in to DNA damage and apoptotic biomarkers is needed