Response of the Erythroid Lineage to Irradiation.
Authors: Wenger Jesse , McGrath Kathleen E. , Koniski Anne , Cacciatori Jenna , Bushnell Timothy P. , Dertinger Stephen D. , Chen Yuhchyau , Palis James 
Published: April 2020

Abstract
Radiation causes acute marrow damage and anemia, however the relative sensitivity of erythroid progenitors and precursors to radiation damage is unknown. While acute anemia elicits a rapid recovery response, the characteristics of erythroid recovery following irradiation have not been well investigated. Mice display a linear response of radiation dose and red cell damage as measured by micronucleated reticulocytes (MN-RET) up to 1-2Gy. Higher radiation doses lead both to decreased MN-RET frequency and to delayed reticulocyte recovery (Dertinger, et al., Mutation Research, in press). To determine why marrow erythroid toxicity differs at low (&lt; 1Gy) versus high (&gt; 2 Gy) radiation doses, we developed a novel method to quantify erythroid precursors using multispectral imaging flow cytometry. C57Bl/6 mice were irradiated either with 0, 1 or 4 Gy and erythroid progenitors (BFU-E, CFU-E) and precursors (ProE, BasoE, PolyE, OrthoE) in the marrow were analyzed between 1 hr and 5 days later. 1 hr following 1Gy radiation, BFU-E and CFU-E were reduced to 63±5% and 13±1% of unirradiated controls, respectively. 24 hrs after 1Gy, there was no significant change in BFU-E, however, a striking recovery of CFU-E (93±21% of controls) was observed. This CFU-E surge was followed by a wave of recovering erythroid precursors in the marrow beginning at 48 hrs and peaking at 72 hrs post radiation, when BasoE, PolyE, and OrthoE reached &gt; 150% of unirradiated controls. These precursors subsequently fell below 70% and 90% of controls at 4 and 5 days, respectively. We also characterized the kinetics of erythroid progenitors and precursors following high-dose radiation. 1 hr after 4Gy, BFU-E and CFU-E were reduced to 10% and 1% of controls, respectively. Each of these erythroid progenitors reached their nadir (1% of controls) at 48 hrs, and began to modestly recover at 72 hrs. By 5 days after 4Gy, BFU-E were only 16±6% of controls, while CFU-E had variably recovered to 127±62%. Erythroid precursors also reached their nadir at 48 hrs following 4Gy with levels of 1–2% of controls, but then recovered to 50–100% by day 5. We conclude that CFU-E are exquisitely sensitive targets of radiation. While BFU-E are more radioresistant, CFU-E recovered more robustly, consistent with the known cycling kinetics of these progenitors. Our results also indicate that the reticulocyte response following low versus high radiation doses is due to rapid CFU-E recovery and the subsequent generation of a wave of erythroid precursors. It is likely that the decrease in MN-RET seen after high-dose radiation is due to the rapid and nearly complete depletion of erythroid precursors leaving few cells able to repair and complete maturation. Finally, we speculate that the oscillation of CFU-E and erythroid precursor populations following 1Gy is due to the operation of feedback regulation. Compared to BFU-E and late-stage erythroid precursors, CFU-E and ProE exhibited a more complex temporal pattern of recovery, suggesting that they may be focal points of this regulation. Examination of marrow progenitor and precursor cell injury and death following acute irradiation will lead to a better understanding of the regulation of hematopoiesis and lay the foundation for interventional studies to mitigate the effects of acute irradiation.