| Objective:Radiation biodosimeter plays a very important role in estimating the absorbed doses of radiation victims and in the research on radiation biological effects. The "golden" biodosimeter, chromosome aberration analysis, is not suit for fast and high-throughput dose estimation. Developing a new fast and simple biodosimeter became the hotspot in radiation biodosimetry. Some studies showed that ionizing radiation could induce the gene expression changes (GEC) in the network of transcriptional regulation pathway, and GEC had the potential to be a biodosimeter. On the basis of preliminary microarray study in our lab, this study was1) to explore the dose-response relationship and time-response range of pig3and gdf15gene that are cultured different times after irradiation;2) to explore the effects of different dose-rate and quality of rays on these two gene expression;3) to explore the effects of other environmental mutagen on these two gene expression.Methods:(1) The dose-effect relationship and time-response range of pig3and gdf15gene exposed to the low-LET radiation:Cell lines AHH-1were irradiated with60Co y-rays, at doses of0,1,3,5,8,10,15and18Gy with dose-rate of1Gy/min. The human lymphoblastoid cells were cultured for8h,12h,24h,48h,72h,5d and7d, respectively after irradiation.(2) The effect of different dose-rate y-ray irradiation on the pig3and gdf15gene expression:Cell lines AHH-1were irradiated with60Co y-rays at doses of0.3,5and8 Gy with dose-rate of0.5Gy/min,1Gy/min or2Gy/min,. The human lymphoblastoid cells were cultured48h after irradiation.(3) The dose-effect relationship of pig3and gdfl5gene exposed to the high-LET radiation:AHH-1cell lines were irradiated with252Cf neutron rays at doses of0,0.4,0.8,1.2and1.6Gy with dose-rate of0.073Gy/min. The human lymphoblastoid cells were cultured48h after irradiation.(4) The effect of other environmental mutagens on pig3and gdfl5gene expression: AHH-1cell lines were treated with H2O2or mitomycin C, combined with or without8Gy60Co y-rays irradiation. The human lymphoblastoid cells were cultured8h,12h and24h respectively after irradiation.Results:(1) At the mRNA level, pig3and gdf15gene expression showed a good dose-effect relationship in a certain dose range (0-10Gy) that cultured8h-72h after irradiation. Their dose-effect relationship could be fitted into a linear or quadratic model with high R" values. While two genes mRNA expression did not show a significant dose-response relationship that cultured5d after irradiation; At7d after irradiation, two genes mRNA expression levels are no statistical difference compared with that of control.(2) At the protein level, pig3and gdfl5gene expression showed a good dose-effect relationship in a certain dose range (0-10Gy) that cultured24h-7d after irradiation whether it is detected by Western blotting or ELISA method. Their dose-effect relationship could be fitted into a linear or quadratic model with high R2values. Because GDF15protein is exocrine protein, we did not detect a significant dose-effect relationship in cell lysates.(3) pig3and gdf15gene expression at mRNA level showed a good dose-effect relationship irradiated with252Cf neutron rays that cultured48h after irradiation and it could be fitted into a linear or quadratic model with high R2values.(4) Whether the transcription level or translation level, pig3and gdf15gene expression did not show statistically significant different at different dose-rates (0.5Gy/min,1Gy/min, and2Gy/min) of3,5, or8Gy.(5) Whether the transcription level or translation level, pig3and gdf15genes expression showed statistically significant difference between the H2O2or mitomycin C treatment group and the control group. And it did not show statistically significant difference between radiation group and irradiation combined with mitogen group.Conclusion:(1) At the transcription level or translation level, detected by real-time PCR, western blot and ELISA, pig3and gdf15gene expression showed a good dose-effect relationship with irradiated dose. This is the fundamental character of radiation biodosimeter.(2) The transcription levels of pig3and gdfl5and translation level of pig3gene expressions showed a good dose-effect relationship irradiated with252Cf neutron rays that cultured48h after irradiation.(3) The transcription levels of pig3and gdf15and translation level of pig3gene expression are not affected by irradiation dose-rate (0.5Gy/min,1Gy/min and2Gy/min) and some environmental mutagens (H2O2and Mitomycin C). |
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