Lymphocyte subsets and their H2AX phosphorylation in response to in vivo irradiation in rats

Abstract

Purpose: The objective of the study was to investigate differences in the radiosensitivity of rat peripheral blood lymphocyte subsets identified by expression of surface clusters of differentiation markers (CD3, CD4, CD8, CD45RA, CD161) after whole-body in vivo gamma-ray irradiation and to assess their individual histone H2AX phosphorylation as an early cell response to irradiation.

Materials and methods: The relative representations of CD45RA B-lymphocytes, CD161 natural killer cells (NK cells), CD3CD4 T-lymphocyte subset and CD3CD8 T-lymphocyte subset in the rat peripheral blood were studied 24–72 hours after irradiation in a dose range of 0–5 Gy. Their intracellular H2AX phosphorylation (γ-H2AX) after 4 Gy and 9 Gy whole-body in vivo irradiation was assessed by multicolour flow cytometry.

Results: We determined the linear dose response of radioresistant CD161 NK cells (24 h), both radiosensitive T-lymphocyte subsets (24 h) and CD45RA B-lymphocytes (72 h) after in vivo irradiation. CD45RA B-lymphocytes showed the highest radiosensitivity and we observed pronounced H2AX phosphorylation which remained expressed in these cells for over 4 h after irradiation.

Conclusion: The combination of the surface immunophenotyping together with intracellular detection of γ-H2AX offers the possibility to assess the absorbed dose of ionizing irradiation with high sensitivity post irradiation and could be successfully applied to biodosimetry.

Keywords::

• radiation
• rats
• lymphocytes
• flow cytometry
• H2AX
• in vivo

Acknowledgements

The authors would like to thank Bc. Lenka Mervartová and Mrs Iveta Tóthová for their excellent technical assistance.

Declaration of interest

The authors report no declaration of interest. The authors alone are responsible for the content and writing of the paper.

This work was supported by the Ministry of Defence of the Czech Republic (The institutional support to a long-term organization development plan 1011, project No. OVUOFVZ200806 and project No. OVUOFVZ200809).