Erythropoietin increases the radioresistance of a clonal hematopoietic progenitor cell line expressing a transgene for the erythropoietin receptor

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Abstract
Erythropoietin (Epo) is a serum glycoprotein growth factor required for the survival, proliferation and differentiation of committed erythroid progenitor cells. In the present study, we sought to determine whether the action of Epo via its receptor is also implicated in the repair of radiation-induced cell damage. Overexpression of the Epo receptor (Epo-R) was achieved as a result of transfection of the 32D cl 3 clonal hematopoietic cell line. These clonal lines allowed us to investigate the effects of Epo on the radiation sensitivity in vitro of a clonal murine hematopoietic progenitor cell line. Low level expression of Epo-R on many hematopoietic cell types was thus circumvented. Ligand binding of Epo resulted in increased radioresistance of 32D cl 3 subclonal lines expressing the Epo-R transgene. The D0 of 32D Epo-R cells at 1.49 Gy/min was 1.33 Gy and n̄ as 1.39. The D0 of parental clonal cell line 32D cl 3 cells at 1.49 Gy/min was 1.36 Gy and n̄ was 1.39. In contrast, at the low dose rate of 0.0595 Gy/min, the D0 of 32D Epo-R cells was 2.0 Gy and n̄ was 1.24, while parental clonal line 32D cl 3 showed a D0 of 1.35 Gy and n̄ was 1.39. The increased radioresistance was statistically significant at low dose rate (p < 0.05). Combined exposure to Epo and interleukin 3 (IL-3) increased proliferation of 32D Epo-R cells but did not induce a detectable further increase in radioresistance. Temporal dissociation between growth factor-activated tyrosine phosphorylation of intracellular substrates, and the radioprotective effect was observed. We searched for possible involvement of an Epo-mediated alteration of cell cycle progression as a cause of radio-protection. A 24-h exposure to Epo alone or in combination with IL-3 did not significantly expand the relative size of the cell population in the relatively radioresistant S phase. The present results indicate that Epo can be considered as one of the group of cytokines that demonstrates a radioprotective effect in vitro. Administration of Epo may have a role in some radiotherapy protocols where specific erythroid hematopoietic toxicity is expected.