Characterization of Radiation-responsive Genes and Transcript Variants under Different Radiation Qualities, Doses and...

Rapid molecular biodosimetry may support radiation exposure assessment during mass casualty or mixed-field radiological incidents, but how gene expression responses vary across radiation qualities and dose rates remains incompletely characterized. Gene expression biodosimetry, including transcript-variant-aware measurements resulting from alternative splicing, may improve sensitivity for determining radiation quality and dose rate. We profiled nine radiation-responsive genes (AEN, APOBEC3H, DDB2, EDA2R, FDXR, PCNA, RPS27L, TRIAP1, ZMAT3) and six FDXR transcript variants using custom 384-well microfluidic RT-qPCR array cards (TaqMan Low Density Array, TLDA) of ex vivo leukopak-derived samples (leukocyte-enriched blood products) from three healthy donors, harvested 24 h after irradiation. Samples received an acute exposure of X rays (0.1-4 Gy), a protracted exposure of X rays (2 Gy at 0.1, 0.01, 0.004 Gy·min-1), and a mixed neutron field exposure with approximately 81% neutrons and 19% concomitant γ-rays (0.1-2 Gy at 0.6-1 Gy·h-1). In a separate experiment, gene expression of leukopaks was concordant with whole blood responses for FDXR and CDKN1A, with FDXR exceeding a 30-fold induction after exposure to 4 Gy of X rays. In the TLDA dataset, all targets were upregulated in a dose-dependent manner. Over the range of 0.1-1 Gy, neutrons elicited higher induction than X rays for APOBEC3H, DDB2, and FDXR, with convergence at 2 Gy as X-ray responses plateaued. Variant-level analysis showed isoform-specific behavior: FDXR-203 and FDXR-206 mRNA isoforms were induced by neutrons (especially at higher doses), while FDXR-202, FDXR-205, and FDXR-217 responded preferentially to X rays and FDXR-204 was moderately upregulated after X-ray irradiation. After protracted X-ray exposure, most genes were up-regulated with minimal dose-rate effects; AEN was the only target that differed significantly between 0.004 and 0.01 Gy·min-1, and FDXR-202/-204/-205 showed a higher fold change at the lowest dose rate. Analysis of dose-response behavior and derived relative biological effectiveness (RBE) values revealed three patterns when comparing neutrons with X rays. First, FDXR, DDB2 and EDA2R were more responsive to neutrons than to X rays at low doses (0.1-1 Gy), with fold changes increasing faster for neutrons. Depending on the gene, neutrons were equivalent to X-ray doses that were approximately 2-4 times higher, consistent with results reported previously for cytogenetic endpoints. Second, APOBEC3H, FDXR-217, RPS27L, PCNA and FDXR-203 showed similar dose-response behavior after exposure to neutrons and X rays, with occasional plateau differences. Third, AEN, TRIAP1 and the remaining assayed FDXR variants showed high variability and were not studied further here. Overall, several genes showed little discrimination across low dose rates, supporting their use for estimating total dose, while others showed clear differences between radiations of different qualities. Taken together, our results support the further development of multi-gene, variant-resolved approaches for radiation exposure assessment in mixed-field scenarios.