Linear Energy Transfer Modulates Radiation-Induced NF-kappa B Activation and Expression of its Downstream Target Genes

Nuclear factor kappaB (NF-kappa B) is a central transcription factor in the immune system and modulates cell survival in response to radiotherapy. Activation of NF-kappa B was shown to be an early step in the cellular response to ultraviolet A (UVA) and ionizing radiation exposure in human cells. NF-kappa B activation by the genotoxic stress-dependent sub-pathway after exposure to different radiation qualities had been evaluated to a very limited extent. In addition, the resulting gene expression profile, which shapes the cellular and tissue response, is unknown. Therefore, in this study the activation of NF-kappa B after exposure to low- and high-linear energy transfer (LET) radiation and the expression of its target genes were analyzed in human embryonic kidney (HEK) cells. The activation of NF-kappa B via canonical and genotoxic stress-induced pathways was visualized by the cell line HEK-pNF-kappa B-d2EGFP/Neo L2 carrying the destabilized enhanced green fluorescent protein (d2EGFP) as reporter. The NF-kappa B-dependent d2EGFP expression after irradiation with X rays and heavy ions was evaluated by flow cytometry. Because of differences in the extent of NF-kappa B activation after irradiation with X rays (significant NF-kappa B activation for doses > 4 Gy) and heavy ions (significant NF-kappa B activation at doses as low as 1 Gy), it was expected that radiation quality (LET) played an important role in the cellular radiation response. In addition, the relative biological effectiveness (RBE) of NF-kappa B activation and reduction of cellular survival were compared for heavy ions having a broad LET range (similar to 0.3-9,674 keV/mu m). Furthermore, the effect of LET on NF-kappa B target gene expression was analyzed by real-time reverse transcriptase quantitative PCR (RT-qPCR). The maximal RBE for NF-kappa B activation and cell killing occurred at an LET value of 80 and 175 keV/mu m, respectively. There was a dose-dependent increase in expression of NF-kappa B target genes NF-kappa B1A and CXCL8. A qPCR array of 84 NF-kappa B target genes revealed that TNF and a set of CXCL genes (CXCL1, CXCL2, CXCL8, CXCL10), CCL2, VCAM1, CD83, NF-kappa B1, NF-kappa B2 and NF-kappa BIA were strongly upregulated after exposure to X rays and neon ions (LET 92 keV/mu m). After heavy-ion irradiations, it was noted that the expression of NF-kappa B target genes such as chemokines and CD83 was highest at an LET value that coincided with the LET resulting in maximal NF-kappa B activation, whereas expression of the NF-kappa B inhibitory gene NFKBIA was induced transiently by all radiation qualities investigated. Taken together, these findings clearly demonstrate that NF-kappa B activation and NF-kappa B-dependent gene expression by heavy ions are highest in the LET range of; similar to 50-200 keV/mu m. The upregulated chemokines and cytokines (CXCL1, CXCL2, CXCL10, CXCL8/IL-8 and TNF) could be important for cell-cell communication among hit as well as nonhit cells (bystander effect). (C) 2018 by Radiation Research Society