Sublethal radiation injury uncovers a functional transition during erythroid maturation

Abstract

Objective Clastogenic injury of the erythroid lineage results in anemia, reticulocytopenia, and transient appearance of micronucleated reticulocytes (MN-RET). However, the MN-RET dose-response in murine models is only linear to 2 Gy total body irradiation (TBI) and paradoxically decreases at higher exposures, suggesting complex radiation effects on erythroid intermediates. To better understand this phenomenon, we investigated the kinetics and apoptotic response of the erythron to sublethal radiation injury. Materials and Methods We analyzed the response to 1 and 4 Gy TBI of erythroid progenitors and precursors using colony assays and imaging flow cytometry (IFC), respectively. We also investigated cell cycling and apoptotic gene expression of the steady-state erythron. Results Following 1 Gy TBI, erythroid progenitors and precursors were partially depleted. In contrast, essentially all bone marrow erythroid progenitors and precursors were lost within two days following 4 Gy irradiation. IFC analysis revealed preferential loss of phenotypic erythroid colony-forming units (CFU-E) and proerythroblasts immediately following sublethal irradiation. Furthermore, these populations underwent radiation-induced apoptosis, without changes in steady-state cellular proliferation, at much higher frequencies than later-stage erythroid precursors. Primary erythroid precursor maturation is associated with marked Bcl-xL upregulation and Bax and Bid down-regulation. Conclusions MN-RET loss following higher sublethal radiation exposures results from rapid depletion of erythroid progenitors and precursors. This injury reveals that CFU-E and proerythroblasts constitute a particularly proapoptotic compartment within the erythron. We conclude that the functional transition of primary proerythroblasts to later-stage erythroid precursors is characterized by a shift from a pro-apoptotic to an anti-apoptotic phenotype.

A Compartmental Model of Mouse Thrombopoiesis and Erythropoiesis to Predict Bone Marrow Toxicity After Internal Irradiation

“…These observations confirmed that BM irradiation directly injured the progenitors whereas the decrease of precursors and mature cells could be explained by the nonrenewal of naturally dying cells, as previously described (19,20). Indeed, a rapid depletion of progenitors was noticed whereas delayed effects were observed for precursors and then for blood cells.…”

Contrasting dynamic responses in vivo of the Bcl-xL and Bim erythropoietic survival pathways

“…These results are consistent with previous reports of increasing Bcl-x L with differentiation in vitro, 20,22 and with the increase in Bcl-x L and decrease in Bid and Bax transcripts with the transition from early-to-late erythroblasts in murine BM. 6 Together with our previous findings of high Fas and FasL in early, but not late, erythroblasts, 5,18,49 a strong pattern emerges of apoptosis-prone early erythroblasts, containing high levels of proapoptotic regulators and only low levels of antiapoptotic proteins, gradually transitioning into apoptosis-resistant late erythroblasts in which antiapoptotic proteins predominate. This underlying pattern explains why high levels of apoptosis are seen in early erythroblasts but not in late erythroblasts during normal fetal and basal adult erythropoiesis in vivo 5,18,19,49 ( Figure 4E), and was recently suggested as responsible for the sensitivity of early erythroblasts to irradiation.…”

“…8,[13][14][15][16] Importantly, it is unknown how multiple survival pathways integrate in vivo to provide a coherent erythropoietic stress response, and whether the large number of survival pathways represents redundancy or functional specialization. The study of these pathways in vivo, now made possible with the advent of flow cytometric techniques, [5][6][7]17 may assist in answering this question.…”

“…4 It was therefore suggested that Epo regulates erythropoietic rate through the number of erythroid progenitors it rescues from apoptosis. 4 Recently, we 5 and others 6,7 made use of cell-surface markers to identify maturation-specific erythroblast subsets directly within freshly isolated mouse hematopoietic tissue, 5 including the Eporesponsive ProE (proerythroblasts, CD71 high Ter119 medium ) and EryA (early basophilic erythroblasts, CD71 high Ter119 high FSC high ). This approach confirmed the central role of apoptosis in the erythropoietic stress response, showing that a substantial fraction of the early erythroblast compartment, particularly splenic ProE and EryA, undergo apoptosis in vivo in the normal basal state.…”

miR-214 protects erythroid cells against oxidative stress by targeting ATF4 and EZH2

“…The increased demand of iron for hemoglobin synthesis in erythroblast results in enhanced formation of labile iron and consequently generation of free radicals, oxidative stress and cytotoxicity [19][20][21]. In addition, erythroid precursors may be also subjected to diverse stresses in bone marrow, such as hypoxia, mechanical stress, radiation, foreign chemicals and environmental toxicants [23][24][25][26]. Thus, an elegant network has evolved to protect erythroid cells during differentiation, including translation control to prevent hemoglobin aggregating by heme-regulated eIF2α kinase (HRI), ATF4 signaling against oxidative stress and Nrf2 signaling-mediated protection, etc [22,27].…”