A higher micronucleus yield in B-versus T-cells after low-dose gamma-irradiation is not linked with defective Ku86 protein.
OBJECTIVE
To elaborate the B-cell micronucleus (MN) response in the low-dose region in detail and to investigate the postulated deficiency in DNA-PK in B-cells.


METHODS
Lymphocytes of five healthy volunteers were irradiated with low LET gamma-rays and high LET fast neutrons with doses ranging between 0.01 and 2 Gy. After post-irradiation incubation, B- and T-cells were isolated via CD3 and CD19 immunomagnetic microbeads. MN were analysed in both subpopulations. To study the underlying mechanism of chromosomal radiosensitivity, cell extracts prepared from purified B- and T-cells were subjected to SDS-electrophoresis and electroblotting using antibodies directed against the DNA-PK repair enzymes Ku70/86 and DNA-PKcs. Activity measurements were performed using the SignaTECT DNA-dependent protein kinase assay. DNA double-strand break (DSB) induction and rejoining was determined using constant-field gel electrophoresis.


RESULTS
For low LET gamma-rays a higher MN yield was observed in B-cells than in T-cells, but only in those samples exposed to doses < 1 Gy. For 1 Gy, the MN yields were comparable and for 2Gy even lower in B-cells compared with T-cells. After high LET neutron irradiation no significant differences in MN yields were observed between both subsets. The results of the DNA-PK experiments demonstrate that there is no difference between T- and B-cells in the basal expression and activity of DNA-PK repair proteins. No differences in DNA DSB induction and rejoining were found between T- and B-cells using constant-field gel electrophoresis.


CONCLUSIONS
From the results, it was concluded that the enhanced chromosomal radiosensitivity in B-cells is restricted to low doses (<1 Gy) of low LET radiation and that the chromosomal behaviour of B-cells to low LET radiation cannot be attributed to aberrant forms of the DNA-PK components. A type of chromosomal induced radioresistance (IRR) may be a possible explanation for the observed effect.
Department of Anatomy, Embryology, Histology and Medical Physics, Ghent University, Belgium.
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Vral
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Thierens
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Bryant
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De Ridder
D000328:Adult; D034961:Antigens, Nuclear; D001402:B-Lymphocytes / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects*; D002467:Cell Nucleus / Q000473:pathology / Q000528:radiation effects; D002469:Cell Separation; D002478:Cells, Cultured; D019457:Chromosome Breakage; D004265:DNA Helicases; D004260:DNA Repair / Q000528:radiation effects; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004307:Dose-Response Relationship, Radiation; D005720:Gamma Rays; D006801:Humans; D018189:Immunomagnetic Separation; D000072200:Ku Autoantigen; D018499:Linear Energy Transfer; D008214:Lymphocytes / Q000166:cytology / Q000528:radiation effects*; D015162:Micronucleus Tests; D008875:Middle Aged; D009502:Neutrons; D009687:Nuclear Proteins / Q000378:metabolism*; D013601:T-Lymphocytes / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects*
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen