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Low Doses of Oxygen Ion Irradiation Cause Acute Damage to Hematopoietic Cells in Mice.

Chang J, Luo Y, Wang Y, Pathak R, Sridharan V, Jones T, Mao XW, Nelson G, Boerma M, Hauer-Jensen M, Zhou D, Shao L - PLoS ONE (2016)

Bottom Line: HZE particles exhibit dense linear tracks of ionization associated with clustered DNA damage and often high relative biological effectiveness (RBE).Furthermore, HPCs and HSCs from irradiated mice exhibited a significant reduction in clonogenic function determined by the colony-forming and cobblestone area-forming cell assays.These acute adverse effects of 16O irradiation on HSCs coincided with an increased production of reactive oxygen species (ROS), enhanced cell cycle entry of quiescent HSCs, and increased DNA damage.

View Article: PubMed Central - PubMed

Affiliation: Division of Radiation Health, Department of Pharmaceutical Sciences, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.

ABSTRACT
One of the major health risks to astronauts is radiation on long-duration space missions. Space radiation from sun and galactic cosmic rays consists primarily of 85% protons, 14% helium nuclei and 1% high-energy high-charge (HZE) particles, such as oxygen (16O), carbon, silicon, and iron ions. HZE particles exhibit dense linear tracks of ionization associated with clustered DNA damage and often high relative biological effectiveness (RBE). Therefore, new knowledge of risks from HZE particle exposures must be obtained. In the present study, we investigated the acute effects of low doses of 16O irradiation on the hematopoietic system. Specifically, we exposed C57BL/6J mice to 0.1, 0.25 and 1.0 Gy whole body 16O (600 MeV/n) irradiation and examined the effects on peripheral blood (PB) cells, and bone marrow (BM) hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) at two weeks after the exposure. The results showed that the numbers of white blood cells, lymphocytes, monocytes, neutrophils and platelets were significantly decreased in PB after exposure to 1.0 Gy, but not to 0.1 or 0.25 Gy. However, both the frequency and number of HPCs and HSCs were reduced in a radiation dose-dependent manner in comparison to un-irradiated controls. Furthermore, HPCs and HSCs from irradiated mice exhibited a significant reduction in clonogenic function determined by the colony-forming and cobblestone area-forming cell assays. These acute adverse effects of 16O irradiation on HSCs coincided with an increased production of reactive oxygen species (ROS), enhanced cell cycle entry of quiescent HSCs, and increased DNA damage. However, none of the 16O exposures induced apoptosis in HSCs. These data suggest that exposure to low doses of 16O irradiation induces acute BM injury in a dose-dependent manner primarily via increasing ROS production, cell cycling, and DNA damage in HSCs. This finding may aid in developing novel strategies in the protection of the hematopoietic system from space radiation.

No MeSH data available.


Related in: MedlinePlus

16O TBI causes persistent increases in DNA damage in HSCs but not in HPCs two weeks after the exposure.(A) Representative analysis of DNA damage measured in Lin- cells by flow cytometry using γH2AX staining in BM HPCs and HSCs from control and 1.0 Gy16O TBI mice. The histograms indicate γH2AX MFI from a representative experiment. (B) The γH2AX MFI in BM HPCs and HSCs after TBI are presented as mean ± SD. (C) Sorted HPCs and HSCs from irradiated and non-irradiated mice were stained with γH2AX antibody. The numbers of foci in each cell were counted and expressed as mean ± SD. (D) The distribution of foci was expressed as the percentages of different numbers of foci in control and irradiated HSCs. The statistical significance for the difference between the control groups and each of irradiated groups is indicated by asterisks. *p<0.05, **p<0.01 by one-way ANOVA analysis.
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pone.0158097.g006: 16O TBI causes persistent increases in DNA damage in HSCs but not in HPCs two weeks after the exposure.(A) Representative analysis of DNA damage measured in Lin- cells by flow cytometry using γH2AX staining in BM HPCs and HSCs from control and 1.0 Gy16O TBI mice. The histograms indicate γH2AX MFI from a representative experiment. (B) The γH2AX MFI in BM HPCs and HSCs after TBI are presented as mean ± SD. (C) Sorted HPCs and HSCs from irradiated and non-irradiated mice were stained with γH2AX antibody. The numbers of foci in each cell were counted and expressed as mean ± SD. (D) The distribution of foci was expressed as the percentages of different numbers of foci in control and irradiated HSCs. The statistical significance for the difference between the control groups and each of irradiated groups is indicated by asterisks. *p<0.05, **p<0.01 by one-way ANOVA analysis.

Mentions: If 16O TBI can induce oxidative stress in HPCs and HSCs, this may also cause DNA damage, known to impair HPC and HSC function. To test this hypothesis, we used flow cytometry to analyze mean fluorescence intensity (MFI) of γH2AX immunostaining in HPCs and HSCs as an indication of DNA double strand breaks (Fig 6A). HPCs from irradiated mice showed no significant change in MFI of γH2AX staining (Fig 6B). However, a significant increase in γH2AX MFI was observed in LSK after all doses of 16O, and in HSCs after the 1.0 Gy dose (Fig 6B, p<0.05-p<0.01). To confirm 16O irradiation-induced DNA damage in HPCs and HSCs, we isolated HPCs and HSCs from irradiated and non-irradiated mice by cell sorting and conducted γH2AX immunostaining. The number of γH2AX foci in each cell was counted under a fluorescent microscope. The data indicated that irradiated HSCs, but not irradiated HPCs, had significantly higher numbers of γH2AX foci per cell compared to non-irradiated HSCs (Fig 6C, p<0.05), which is consistent with the data from MFI of γH2AX by flow cytometry. Moreover, we further analyzed the foci distribution in HSCs. The results showed that more than 60% of HSCs from irradiated and non-irradiated mice did not have γH2AX foci. About 15–17% of HSCs contained more than two γH2AX foci in each HSC from irradiated mice while only 9% of normal HSCs had two foci per cell (Fig 6D). Therefore, these data demonstrate that 16O irradiation induces DNA damage in irradiated LSK and HSCs, which may contribute to 16O irradiation-induced acute hematopoietic damage.


Low Doses of Oxygen Ion Irradiation Cause Acute Damage to Hematopoietic Cells in Mice.

Chang J, Luo Y, Wang Y, Pathak R, Sridharan V, Jones T, Mao XW, Nelson G, Boerma M, Hauer-Jensen M, Zhou D, Shao L - PLoS ONE (2016)

16O TBI causes persistent increases in DNA damage in HSCs but not in HPCs two weeks after the exposure.(A) Representative analysis of DNA damage measured in Lin- cells by flow cytometry using γH2AX staining in BM HPCs and HSCs from control and 1.0 Gy16O TBI mice. The histograms indicate γH2AX MFI from a representative experiment. (B) The γH2AX MFI in BM HPCs and HSCs after TBI are presented as mean ± SD. (C) Sorted HPCs and HSCs from irradiated and non-irradiated mice were stained with γH2AX antibody. The numbers of foci in each cell were counted and expressed as mean ± SD. (D) The distribution of foci was expressed as the percentages of different numbers of foci in control and irradiated HSCs. The statistical significance for the difference between the control groups and each of irradiated groups is indicated by asterisks. *p<0.05, **p<0.01 by one-way ANOVA analysis.
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Related In: Results  -  Collection

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pone.0158097.g006: 16O TBI causes persistent increases in DNA damage in HSCs but not in HPCs two weeks after the exposure.(A) Representative analysis of DNA damage measured in Lin- cells by flow cytometry using γH2AX staining in BM HPCs and HSCs from control and 1.0 Gy16O TBI mice. The histograms indicate γH2AX MFI from a representative experiment. (B) The γH2AX MFI in BM HPCs and HSCs after TBI are presented as mean ± SD. (C) Sorted HPCs and HSCs from irradiated and non-irradiated mice were stained with γH2AX antibody. The numbers of foci in each cell were counted and expressed as mean ± SD. (D) The distribution of foci was expressed as the percentages of different numbers of foci in control and irradiated HSCs. The statistical significance for the difference between the control groups and each of irradiated groups is indicated by asterisks. *p<0.05, **p<0.01 by one-way ANOVA analysis.
Mentions: If 16O TBI can induce oxidative stress in HPCs and HSCs, this may also cause DNA damage, known to impair HPC and HSC function. To test this hypothesis, we used flow cytometry to analyze mean fluorescence intensity (MFI) of γH2AX immunostaining in HPCs and HSCs as an indication of DNA double strand breaks (Fig 6A). HPCs from irradiated mice showed no significant change in MFI of γH2AX staining (Fig 6B). However, a significant increase in γH2AX MFI was observed in LSK after all doses of 16O, and in HSCs after the 1.0 Gy dose (Fig 6B, p<0.05-p<0.01). To confirm 16O irradiation-induced DNA damage in HPCs and HSCs, we isolated HPCs and HSCs from irradiated and non-irradiated mice by cell sorting and conducted γH2AX immunostaining. The number of γH2AX foci in each cell was counted under a fluorescent microscope. The data indicated that irradiated HSCs, but not irradiated HPCs, had significantly higher numbers of γH2AX foci per cell compared to non-irradiated HSCs (Fig 6C, p<0.05), which is consistent with the data from MFI of γH2AX by flow cytometry. Moreover, we further analyzed the foci distribution in HSCs. The results showed that more than 60% of HSCs from irradiated and non-irradiated mice did not have γH2AX foci. About 15–17% of HSCs contained more than two γH2AX foci in each HSC from irradiated mice while only 9% of normal HSCs had two foci per cell (Fig 6D). Therefore, these data demonstrate that 16O irradiation induces DNA damage in irradiated LSK and HSCs, which may contribute to 16O irradiation-induced acute hematopoietic damage.

Bottom Line: HZE particles exhibit dense linear tracks of ionization associated with clustered DNA damage and often high relative biological effectiveness (RBE).Furthermore, HPCs and HSCs from irradiated mice exhibited a significant reduction in clonogenic function determined by the colony-forming and cobblestone area-forming cell assays.These acute adverse effects of 16O irradiation on HSCs coincided with an increased production of reactive oxygen species (ROS), enhanced cell cycle entry of quiescent HSCs, and increased DNA damage.

View Article: PubMed Central - PubMed

Affiliation: Division of Radiation Health, Department of Pharmaceutical Sciences, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.

ABSTRACT
One of the major health risks to astronauts is radiation on long-duration space missions. Space radiation from sun and galactic cosmic rays consists primarily of 85% protons, 14% helium nuclei and 1% high-energy high-charge (HZE) particles, such as oxygen (16O), carbon, silicon, and iron ions. HZE particles exhibit dense linear tracks of ionization associated with clustered DNA damage and often high relative biological effectiveness (RBE). Therefore, new knowledge of risks from HZE particle exposures must be obtained. In the present study, we investigated the acute effects of low doses of 16O irradiation on the hematopoietic system. Specifically, we exposed C57BL/6J mice to 0.1, 0.25 and 1.0 Gy whole body 16O (600 MeV/n) irradiation and examined the effects on peripheral blood (PB) cells, and bone marrow (BM) hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) at two weeks after the exposure. The results showed that the numbers of white blood cells, lymphocytes, monocytes, neutrophils and platelets were significantly decreased in PB after exposure to 1.0 Gy, but not to 0.1 or 0.25 Gy. However, both the frequency and number of HPCs and HSCs were reduced in a radiation dose-dependent manner in comparison to un-irradiated controls. Furthermore, HPCs and HSCs from irradiated mice exhibited a significant reduction in clonogenic function determined by the colony-forming and cobblestone area-forming cell assays. These acute adverse effects of 16O irradiation on HSCs coincided with an increased production of reactive oxygen species (ROS), enhanced cell cycle entry of quiescent HSCs, and increased DNA damage. However, none of the 16O exposures induced apoptosis in HSCs. These data suggest that exposure to low doses of 16O irradiation induces acute BM injury in a dose-dependent manner primarily via increasing ROS production, cell cycling, and DNA damage in HSCs. This finding may aid in developing novel strategies in the protection of the hematopoietic system from space radiation.

No MeSH data available.


Related in: MedlinePlus