Effects of culturing technique on human peripheral blood lymphocytes response to proton and X-ray radiation

Abstract

Purpose: The main aim of this study was to comparatively investigate the effects of culturing methods on the response of human peripheral blood lymphocytes to irradiation exposure.

Materials and methods: Whole blood and isolated lymphocytes were ex vivo exposed to two radiation sources (60 MeV proton or 250 kV X-ray radiation) with different doses (0.3, 0.5, 0.75, 1.0, 1.5, 2.0, 2.5, 3.0, and 4.0 Gy), and genotoxic markers were subsequently assayed. The observed effects were compared as dose–response relationships using two end points (CBMN and PCC tests) and different biomarkers (NDI, PCC index, MNi frequency and excess PCC fragments).

Results and conclusions: The results showed different effects of the culturing techniques on the response of human peripheral blood lymphocytes to radiation. The MNi frequency and excess PCC fragments were significantly higher when lymphocytes were cultured after being isolated. After irradiation, no differences were seen in the NDI between the lymphocytes of the two culturing techniques; however, there were differences in the PPC index. When planning or performing cytogenetic studies, the possibility of such effects and their potential to impact the variability of the results of human biomonitoring studies should be considered important and taken into account.

Keywords:

• Human peripheral blood lymphocytes
• protons
• isolation process
• whole blood samples
• premature chromosome condensation (PCC) test
• cytokinesis-block micronucleus (CBMN) assay

View correction statement:

Correction

Acknowledgments

These investigations were carried out as part of an extended examination of the proton beam at IFJ PAN by cytogenetic and molecular methods and were partially supported by Grant DEC-2013/09/D/NZ7/00324 from the National Science Centre, Poland and the project co-funded by the Malopolska Regional Operational Program Measure 5.1 Krakow Metropolitan Area as an important hub of the European Research Area for 2007-2013 (access to laboratory equipment). Authors gratefully acknowledge the donor for providing blood samples, technical staff for irradiation and dosimetry, “Diagnostics” company in Kraków, Poland for help in blood samples collection. Authors would like to cordially thank Agnieszka Panek, PhD, Iwona Możdżeń, MSc, Jolanta Adamczyk, Janusz Swakoń, PhD and Renata Kopeć, PhD for their help in the laboratory work and irradiation performance and preparadness.

Author contributions

Justyna Miszczyk conceived an idea and collaborative project, together with Kamila Rawojć carried out the experiments and measurements, wrote the manuscript as well as the statistics, tables and figures. Both have given approval to the final version of the manuscript.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Notes on contributors

Justyna Miszczyk

Justyna Miszczyk, PhD, has an extent experience in radiobiology. Dr. Miszczyk's research focuses on effects of normal tissue damage induced by different types of radiation.

Kamila Rawojć

Kamila Rawojć, PhD, Trained medical physicist with experience in radiotherapy planning and nuclear medicine imaging. Dr. Rawojc's research is focused protonl radiobiology.