Alpha-tocopherol succinate- and AMD3100-mobilized progenitors mitigate radiation combined injury in mice

Abstract

The purpose of this study was to elucidate the role of alpha-tocopherol succinate (TS)- and AMD3100-mobilized progenitors in mitigating combined injury associated with acute radiation exposure in combination with secondary physical wounding. CD2F1 mice were exposed to high doses of cobalt-60 gamma-radiation and then transfused intravenously with 5 million peripheral blood mononuclear cells (PBMCs) from TS- and AMD3100-injected mice after irradiation. Within 1 h after irradiation, mice were exposed to secondary wounding. Mice were observed for 30 d after irradiation and cytokine analysis was conducted by multiplex Luminex assay at various time-points after irradiation and wounding. Our results initially demonstrated that transfusion of TS-mobilized progenitors from normal mice enhanced survival of acutely irradiated mice exposed 24 h prior to transfusion to supralethal doses (11.5-12.5 Gy) of (60)Co gamma-radiation. Subsequently, comparable transfusions of TS-mobilized progenitors were shown to significantly mitigate severe combined injuries in acutely irradiated mice. TS administered 24 h before irradiation was able to protect mice against combined injury as well. Cytokine results demonstrated that wounding modulates irradiation-induced cytokines. This study further supports the conclusion that the infusion of TS-mobilized progenitor-containing PBMCs acts as a bridging therapy in radiation-combined-injury mice. We suggest that this novel bridging therapeutic approach involving the infusion of TS-mobilized hematopoietic progenitors following acute radiation exposure or combined injury might be applicable to humans.

Keywords: cytokines; gamma-radiation; hematopoietic progenitor cells; mice; transfusion.

Fig. 1.

Diagrammatic representation of experimental model for evaluating TS mobilized progenitors as a radio-mitigator and countermeasure against combined injury (radiation plus wound). Donor mice received TS and AMD3100 72 h and 1 h, respectively, before blood collection. PBMCs were transfused into irradiated mice 24 h after irradiation. Combined injury mice received wound 1 h after irradiation.

Fig. 2.

Efficacy of TS-mobilized progenitors as a radio-mitigator against different doses of ⁶⁰Co γ-radiation in mice. Five million PBMCs from TS-injected mice were transfused to each mouse 24 h after irradiation with 11.5 (n = 16), 12.0 (n = 16), and 12.5 (n = 16) Gy. Survival was monitored for 30 d. The table shows the statistical significance of the differences between vehicle control and TS-mobilized, progenitor-administered groups at each radiation dose on Days 10, 15 and 30 (ns denotes not significant).

Fig. 3.

Effect of TS-mobilized progenitors on survival of mice exposed to combined injury (irradiation and wounding). TS was administered to CD2F1 donor mice (n = 72), and blood was collected from donor mice 72 h after TS injection for isolation of PBMCS. Four groups of mice (n = 16 in each group) were irradiated either with 9.2 or 11 Gy. Two groups in each set were wounded within 1 h following radiation exposure. Five million PBMCs obtained from donor mice were injected 24 h after irradiation into each mouse of the group that had been irradiated and wounded, and also into the group treated with radiation alone. Untreated group mice did not receive cell transfusion. Survival was monitored for 30 d. In an experiment using 9.2 Gy, the TS-mobilized, PBMC-treated group had significantly higher survival compared with controls (†combined injury, P < 0.05, or *irradiated only, P < 0.001). In an experiment using 11 Gy, TS-mobilized, PBMC-treated mice had a higher number of survivors compared with its control (*P < 0.01), but the difference in the wounded groups was not significant.

Fig. 4.

Effect of TS treatment on the survival of mice with combined injury (exposed to 9.2 Gy ⁶⁰Co γ-radiation and wounded). TS or vehicle was administered to mice (n = 16) 24 h before radiation exposure, and wounded groups were given a wound within 1 h following irradiation. Survival was monitored for 30 d. Mice treated with TS were protected significantly (*P < 0.001) compared with vehicle control. In combined-injury groups, TS treatment increased the number of survivors over vehicle-treated mice (^†P < 0.01).

Fig. 5.

Time-course of cytokine levels in mice exposed to 8-Gy radiation and treated with TS-mobilized progenitors. CD2F1 donor mice received TS and blood was collected from donor mice for isolation of PBMCs as stated above. Two groups of mice (n = 8) were irradiated and were subsequently injected with either five million TS-mobilized PBMCs obtained from donor mice or vehicle. Blood samples for cytokine analysis were collected 3, 5, 8, 12, 15, 20 and 30 d after irradiation for serum cytokine analysis. *Denotes statistical significance of P < 0.05 between TS PBMCs and vehicle. ^‡P = 0.058.

Fig. 6.

Time-course of cytokine levels in mice receiving radiation exposure and wounding, and treatment of TS-mobilized progenitors. PBMCs were collected from TS-injected mice as described above. Four groups of mice (n = 8) were irradiated with 11 Gy. Two groups were wounded 1 h following radiation exposure. Five million PBMCs from donor mice were injected into one of each group of irradiated and wounded mice, and irradiated only mice. Blood samples were collected from irradiated mice 3 and 7 d after irradiation and analyzed for serum cytokines. There were not enough survivors in the untreated wounded group 7 d post-irradiation for cytokine analysis. *Denotes statistical significance of P < 0.05 between TS PMBCs and TS PBMCs + Wounded, ^#P < 0.01 between untreated and untreated + wounded.

Fig. 7.

Time-course of cytokine levels in mice receiving radiation exposure and wounding, and treatment of TS. TS or vehicle was injected to mice 24 h prior to irradiation. Mice (n = 8) were irradiated (9.2 Gy, dose rate 0.6 Gy/min), and one group of each treatment was wounded 1 h following radiation exposure. Blood samples were collected 3 and 7 d after irradiation for serum cytokine analysis. *Denotes statistical significance of P < 0.01 between TS and vehicle, ^†P < 0.001 between TS + wounded and vehicle + wounded, ^#P < 0.01 between untreated and untreated + wounded.