The response of human mesenchymal stem cells to internal exposure to tritium β-rays

Abstract

There is no doubt that estimating the exposure risk of external and internal low-dose radiation is an imperative issue in radiobiological study. Human mesenchymal stem cells (hMSCs) are multipotent and self-renewing, supporting the regeneration of damaged tissue, including tissue damaged by radiation. However, the responses of hMSCs to internal exposure to radionuclides are still insufficiently understood. In order to evaluate the adverse effects produced by internal exposure to tritiated water (HTO) at a low dose, hMSCs were exposed to 2 × 107 Bq/ml HTO, and the biological effects after the exposure were examined. Apoptosis and DNA double-strand breaks (DSBs) were assayed to analyze the cellular response to the damage induced by HTO. Slight enhancement of apoptosis was found after treatment, except at the dose of 9 mGy. The number of DSBs at 24 h post-irradiation showed that the DNA damage was able to be efficiently repaired by the hMSCs. Moreover, the increasing proportion of the cell population in S phase proved that the persistence of residual γH2AX foci at lower concentrations of HTO was attributable to the secondary production of DSBs in DNA replication. Our work adds to the available data, helping us understand the risk of stem cell transformation due to internal exposure and its correlation with low-dose radiation-induced carcinogenesis.

Keywords: internal exposure; low-dose radiation; stem cell; tritiated water.

Fig. 1.

Apoptosis of cells exposed to 2 × 10⁷ Bq/ml HTO with various doses. Cells were labeled with Annexin V-FITC and PI at 3 h and 24 h post HTO treatment; more than 10 000 cells were collected and the fluorescence was analyzed with flow cytometry. Data were represented as mean ± SD. For comparisons between irradiated groups and sham-irradiated group, values of P < 0.05 are referred to as significant.

Fig. 2.

Average γH2AX foci number per cell produced by 2 × 10⁷ Bq/ml HTO. Before the cells were fixed, they were washed out with fresh medium twice. Then cells harvested at 3 h or 24 h post radiation were assessed via immunofluorescence. Data are represented as mean ± SD. For comparisons between irradiated groups and sham-irradiated group, values of P < 0.05 are referred to as significant.

Fig. 3.

Detection of γH2AX foci in cells exposed to HTO with various levels of radioactivity but the same culture time. (A) Induced γH2AX foci per cell counted at 3 h or 24 h post incubation in HTO with various levels of radioactivity; (B) statistical data for γH2AX foci detected at 3 h or 24 h; (C–H) representative picture of γH2AX foci distribution in damaged cells at 3 h or 24 h post HTO treatment with various levels of radioactivity but the same culture time. Scale bar: 10 μm.

Fig. 4.

Analysis of cell cycle distribution after exposure to various levels of radioactivity HTO. (A) Cells were treated with 2 × 10⁶ Bq/ml and 2 × 10⁵ Bq/ml HTO, separately, and the distribution of cells in the cell cycle was measured with flow cytometry at 3 h post irradiation; (B) the proportion of cells at different cell cycle phases was measured at 24 h post irradiation.