Measurement of mutant frequency in T-cell receptor (TCR) gene by flow cytometry after X-irradiation on EL-4 mice lymphoma cells.

Abstract

It is well known that somatic mutations are induced by ionizing irradiation. We have previously reported the measurement of mutant frequency (MF) on the T-cell receptor (TCR) gene in mouse T-lymphocytes after irradiation by flow cytometry. In this study, we developed an in vitro system using murine EL-4 lymphoma cells and observed frequency of cells defective in TCR gene expression after exposure to ionizing irradiation. EL-4 cells were stained with fluorescein-labeled anti-CD4 and phycoerythrin-labeled anti-CD3 antibodies. They were analyzed with a flow cytometer to detect mutant EL-4 cells lacking surface expression of TCR/CD3 complexes which showed CD3-, CD4+ due to a somatic mutation at the TCR genes. Mutant cells could be observed at 2 days after 3 Gy irradiation. MF of EL-4 cells was 6.7x10(-4) for 0 Gy and the value increased to the maximum level of 39x10(-4) between 4 and 8 days after 3 Gy irradiation and these data were found to be best fitted by a linear-quadratic dose-response model. After the peak value the TCR MF gradually decreased with a half-life of approximately 3.2 days. We also examined the hprt mutant frequencies at seven days after irradiation and the cytokinesis-blocked micronucleus frequency at 20 hrs after irradiation. The frequencies of hprt mutation and micronuclei were found to be best fitted by a linear-quadratic dose-response model and a linear dose-response model, respectively. The method to detect mutation on TCR gene is quick and easy in comparison with other methods and is considered useful for the mutagenicity test.

• Authors: Naoki Kunugita, Nan Mei, Tatjana Goncharova, Toshiyuki Norimura
• Journal: The Journal of toxicological sciences (J Toxicol Sci) Vol. 32 Issue 4 Pg. 377-86 (Oct 2007) ISSN: 0388-1350 [Print] Japan
• PMID: 17965552 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Receptors, Antigen, T-Cell
• Hypoxanthine Phosphoribosyltransferase

Topics

• Animals
• Cell Line, Tumor
• Cell Survival (radiation effects)
• Dose-Response Relationship, Radiation
• Flow Cytometry
• Gene Expression (radiation effects)
• Hypoxanthine Phosphoribosyltransferase (genetics)
• Mice
• Micronucleus Tests
• Mutation
• Receptors, Antigen, T-Cell (genetics)
• T-Lymphocytes (metabolism, radiation effects)
• X-Rays (adverse effects)