The quality of the ionizing radiation (IR) can be described in terms of its nature, photons or particles, and their corresponding energies. The energy is classified in terms of High or Low linear energy transfer that will produce a different distribution of DNA damage and other molecules in the cell either by direct action or indirect action. Indirect action leads to the production of reactive oxygen species (ROS) modifying nucleotides in DNA or free dNTPs. 8-oxo-dGTP is formed through ROS endogenously when there is an imbalance between the antioxidants defence systems and the production of ROS levels in favour of ROS, leading to an oxidative stress condition. Organisms, organs, and cell types show different degrees of radiosensitivity, and this thesis aimed to investigate the underlying mechanisms of IR induced oxidative stress and its relation with radiosensitivity.

In previous studies, we identified proteins involved in radiation response with a focus on low dose radiation response. Cell models were established in which the expression of some protein/s was downregulated by knocking down/out using CRISPR/Cas9 or shRNA technology. The knockdown or knockout cells were exposed to different doses at low dose rates (LDR) or high dose rate (HDR) to investigate the role of these genes/proteins for survival (radiosensitivity), mutation induction, stress response, differentiation, etc. and they were subjected to further studies in this thesis.

Publication I, cell lines with hMTH1, and MYH knockdown were established and exposed to 0.5 and 1 Gy administered at different dose rates. We found that LDR induces significantly increased levels of extracellular 8-oxo-dG compared to HDR. We also found that hMTH1 and MYH play together an important role in the protection of cells against ROS-induced mutagenicity.

Publication II, the role of NRF2 was investigated for the radiosensitivity of glioblastoma cancer stem cells (CSCs). The neutrosphere cells from the U87MG cell line were irradiated with three different radiation qualities. The results show that cells exposed to LDR produce significantly higher levels of extracellular 8-oxo-dG compared to HDR and carbon ion irradiated cells. Lower proliferation, self-renewal, and neurosphere formation were observed in both LDR and HDR irradiated NRF2-knockdown cells as compared with the wild type. The results show that NRF2 plays an important role in the radiosensitivity of neurosphere cells isolated from the U87MG cell line.

Publication III, we examined the relation between 8-oxo-dG levels and the outcome of radiotherapy and chemotherapy in gastrointestinal cancer patients. The results showed that patients with improved treatment outcomes (responders), had lower levels of the stress marker extracellular 8-oxo-dG before the start of the treatment and the levels were increased 2 weeks after completing the treatment.

Publication IV, mice were whole-body irradiated with different doses administered at LDR and HDR. Three hours or three weeks after exposure, the immune cell populations in the spleens were phenotyped. The effects of dose, dose rate, and time after exposure and interaction between them were investigated to check which of the factors had the main effect on the change of immune cell populations. The results indicate that there was a pro-inflammatory short-term effect at high doses for both HDR and LDR. The results also indicate a pro-inflammatory effect of low doses of radiation three weeks after exposure.

Assessment of human health risks from exposure to ionizing radiation (IR) is mainly based on the extrapolation of results from epidemiological studies on populations exposed to relatively high doses and often at high dose rates (HDR). Risk estimates after exposure to low doses and in particular at low dose rates (LDR) remain controversial due to a lack of epidemiological evidence. Therefore, high priority is given to strengthening the evidence on which risk assessments can be based for low doses and LDR. It is known that the cytotoxicity of radiation decreases by decreasing dose rate. Less is known about the effects of LDR on mutation rates and premature senescence compared to HDR. We established 2 cell lines with low expression of two proteins, MTH1 or MYH, both involved in the protection of cells from mutation induction by reactive oxygen species (ROS). The cells were exposed to different doses at different dose rates, and the levels of mutation were studied. The results showed a possible dose-rate threshold for mutations for the MTH1/MYH double knockdown cells.

Next, we studied the effect of dose rate on adaptive response (AR). AR is defined as the ability of a low dose of ionizing radiation to induce enhanced resistance in cells subsequently exposed to a high dose. We established dose response relations for survival and mutations for MCF-10A cells exposed/non-exposed to an adaptive dose of 50 mGy at different dose rates, followed by exposure to different high doses. We found no protective effect of 50 mGy on survival. However, we observed that 50 mGy the adaptive dose reduced the mutation frequency induced by 1 Gy challenging dose. The protection level was higher when 50 mGy was delivered at LDR.

A significant amount of data suggests that oxidative stress, induced for example by LDR, can contribute to senescence. We cultured VH10 cells, beginning with passage 13, during chronic LDR exposure. The cells were passaged every week for 6 weeks until they stopped proliferating due to premature senescence at passage 19. Passage 8 VH10 cells were cultured correspondingly but without irradiation until they stopped proliferating at passage 23 in response to replicative senescence. The DNA repair kinetics and the levels of DNA damage that were localized in the telomeres of young, middle-aged, premature senescent and replicative senescent cells were investigated. The young cells repaired DSB significantly faster than the senescent cells; premature and replicative senescent cells accumulated more DNA damage in the telomeres; and as compared to middle-aged cells, young cells cope with oxidative stress of chronic irradiation more effectively.

The transgenerational effects of IR were studied in Drosophila embryos. The exposed embryos were followed up for abnormality during embryogenesis until adult stage and up to 12 generations. We found that radiation induced an A5pig- phenotype (depigmented area in the A5 segment of the male body) that was transmitted up to 12 generations. This phenomenon did not follow the Mendelian inheritance model, which indicates the influence of mechanisms other than mutagenesis e.g. epigenetic mechanism.

We showed that; LDR is less cytotoxic than HDR but both induce equal levels of mutation per unit dose; LDR induces premature senescence; LDR may be more effective than HDR in inducing adaptive response; and LDR and HDR exposure of Drosophila embryos can induce an abnormal phenotype that can be transmitted through generations.