BACKGROUND: Biodosimetry with persistent cytogenetic indicators in peripheral blood lymphocytes (PBLs) plays crucial role in regulatory/medical management of individuals overexposed to radiation. Conventional methods require ∼48 h culture and have limited dose range (0.1-5Gy) applications due to checkpoint arrest/poor stimulation. G0-Phase Premature chromosome condensation (G0-PCC) allows chromosome aberration analysis within hours after blood collection. Due to high skill demand, applications of G0-PCC were not very well explored and being re-visited worldwide. Among all aberrations, analysis of excess chromosomal fragments is quickest. Radiation dose response curve for the fragments has been reported. PURPOSE: In present study, excess fragment analysis has been addressed in detail, in addition to validation of radiation dose response curve, gender variation in the response, dose dependent repair kinetics, minimum detection limit (MDL), duration and accuracy of final dose estimation with 5blindfolded, ex vivo irradiated samples have been studied. In extension, feasibility of multiparametric dosimetry with Fluorescent in situ hybridization (FISH) based endpoints were qualitatively explored. MATERIAL AND METHODS: PBLs were exposed to Gamma-Radiation and G0-PCC was performed at different time points. Decay kinetics and dose response curve were established. Gender Variation of the frequency of the fragments was assessed at 0, 2 and 4 Gy. FISH was performed with G0-PCC applying centromere probe, whole chromosome paints, multi-color FISH and multi-color banding probes. RESULTS: Radiation response curve for fragments was found to be linear (Slope 1.09 ± 0.031 Gy-1). Background frequency as well as dose response did not show significant gender bias. Based on variation in background frequency of fragments MDL was calculated to be ∼0.3 Gy. Kinetics of fragment tested at 0, 4, 8, 16 and 24 h showed exponential decay pattern from 0 to 8 h and without further decay. Final dose estimation of five samples was completed within 13 man-hours. Dicentric chromosomes, translocations, insertions and breaks were identifiable in combination with centromere FISH and WCP. Advanced methods employing multicolor FISH and multi-color banding were also demonstrated with PCC spreads. CONCLUSION: G0-PCC, can be useful tool for high dose biodosimetry with quick assessment of fragment frequency. Further, it holds potential for multi-parametric dosimetry in combination with FISH.
BACKGROUND: Biodosimetry with persistent cytogenetic indicators in peripheral blood lymphocytes (PBLs) plays crucial role in regulatory/medical management of individuals overexposed to radiation. Conventional methods require ∼48 h culture and have limited dose range (0.1-5Gy) applications due to checkpoint arrest/poor stimulation. G0-Phase Premature chromosome condensation (G0-PCC) allows chromosome aberration analysis within hours after blood collection. Due to high skill demand, applications of G0-PCC were not very well explored and being re-visited worldwide. Among all aberrations, analysis of excess chromosomal fragments is quickest. Radiation dose response curve for the fragments has been reported. PURPOSE: In present study, excess fragment analysis has been addressed in detail, in addition to validation of radiation dose response curve, gender variation in the response, dose dependent repair kinetics, minimum detection limit (MDL), duration and accuracy of final dose estimation with 5blindfolded, ex vivo irradiated samples have been studied. In extension, feasibility of multiparametric dosimetry with Fluorescent in situ hybridization (FISH) based endpoints were qualitatively explored. MATERIAL AND METHODS: PBLs were exposed to Gamma-Radiation and G0-PCC was performed at different time points. Decay kinetics and dose response curve were established. Gender Variation of the frequency of the fragments was assessed at 0, 2 and 4 Gy. FISH was performed with G0-PCC applying centromere probe, whole chromosome paints, multi-color FISH and multi-color banding probes. RESULTS: Radiation response curve for fragments was found to be linear (Slope 1.09 ± 0.031 Gy-1). Background frequency as well as dose response did not show significant gender bias. Based on variation in background frequency of fragments MDL was calculated to be ∼0.3 Gy. Kinetics of fragment tested at 0, 4, 8, 16 and 24 h showed exponential decay pattern from 0 to 8 h and without further decay. Final dose estimation of five samples was completed within 13 man-hours. Dicentric chromosomes, translocations, insertions and breaks were identifiable in combination with centromere FISH and WCP. Advanced methods employing multicolor FISH and multi-color banding were also demonstrated with PCC spreads. CONCLUSION: G0-PCC, can be useful tool for high dose biodosimetry with quick assessment of fragment frequency. Further, it holds potential for multi-parametric dosimetry in combination with FISH.
Authors: Elena Obrador; Rosario Salvador-Palmer; Juan I Villaescusa; Eduardo Gallego; Blanca Pellicer; José M Estrela; Alegría Montoro Journal: Antioxidants (Basel) Date: 2022-05-31