| The rapid development of nuclear energy has profoundly influenced the modern life, however, the side adverse effects such as the radiation damagecauses great concern. Radiation protection becomes more and more important and the priority in injured rescue is estimation the dose of radiation exposal. How to determine the dose of exposure and the severity of the injury more rapidly and reliably is very much pivotal. Cellchromosome distortion analysis is the gold standard method for the assessment of dose dependent cause of the radiation damage. Most of the radiation-induced micronuclei are acentric and the chromosome as well as the ratio of micronuclei increase proportionally depending the increase in the radiation dose. CB micronucleus method has solved the problem of muclear loss in the cells in the rasditional normal cell culture method, making the micronuclei counting more accurate and increase the accuracy of radiation exposure dos determination.Radiation causes DNA damage in cells which leads to the differential gene expression of cell cycle related genes. The cell cycle related genes participate in the repair of DNA damaged by the over exposure of radiation and play an important role in cell cycle regulation and maintain genomic stability.Objective:To screen the expression of radiation response genes as well as to observe the DNA damage induced by various doses of radiation and the differential expression of cell cycle related genes in the X-ray irradiated human peripheral blood lymphocytes as well as to establish a dose-response relationship. Compare to the tradition biological dosimeter results, give assessment of this new method, decide new biomarkers of radiation injury to establish experiment basis of minimally invasive, rapidly detection method, to meet the needs of radiation biological dose in time estimation in the nuclear and radiation medical during emergency conditions.Method:The normal human blood cells were exposed to various doses of X-ray irradiation such as 0, 1, 2, 3, 4, 5 Gray for 4h and 24h respectively. We have used CB micronucleus method to determine the change of micronucleus ratio. Moreover , we have used quantitative Real-Time qPCR to measure the expression of Cdkn1a, Gadd45a, ATM, Egr-1, CyclinB1 and CyclinD1. Statistical analysis with regression analysis to obtain dose-response relationship of the expression of various genes has been established. Finally, we have compared the expression of these genes with the results obtained from CB micronucleus method and assessed the outcome of the radiation induced changes in various groups based on the dose as well as the time.Result:1. The effect of the ionizing radiation on micronucleus ratio in human peripheral blood lymphocytes. The CB micronucleus method indicates that after 4 and 24 hours of X-ray irradiation 0～5 Gray, the ratio of micronuclei has been increased significantly (p<0.05). The linear equation of dose-response relationship can be obtained from the regression analysis with the increase in the radiation dose.2. Ionizing radiation effect on Cell Cycle related gene expression in the human peripheral blood lymphocytes.Real-Time qPCR shows that Cdkn1a gene expression of peripheral blood lymphocytes increased significantly after 0～5 Gray of X-ray irradiation in both time points (4h and 24h), and the increase is dose-dependent. Although the peak is higher in 4Gy than 5Gy group, there is no significant difference between them (p>0.05). In general, the results show that there is indeed a linear relationship exists between the radiation dose-response. Similary , the Gadd45a gene expression of human peripheral blood lymphocytes also increased significantly (p<0.05) after 0～5 Gray X-ray irradiation in both time points (4h and 24h), and is does-dependent. The comparision of different time points show that 4 hour group has higher expression of Gadd45a than 24 hour group. However, X-ray irradiation upregulated the Gadd45a gene expression, and there is a linera relationship between the dose and the time of exposure.3. Expression of CyclinD1 gene in human peripheral blood lymphocytes shows that, at 4 h time point, it increased significantly, and there is a linear relationship between the dose and the expression, and no significant change at 24h time point after 0～5 Gray X-ray irradiation compared with the control group..4. For the genes of ATM, Egr-1, CyclinB1 expression in human peripheral blood lymphocytes, there are no significant difference compared to the control group after 0～5 Gy X-ray irradiation at both time points (4h and 24h), and the expression is not dose dependent. Hence , a linear equation has not been obtained through regression analysis which indicates that there is no linear relationship between the doses.Conclusion:1. X-ray irradiation causes the increase in the ratio of micronuclei in human peripheral blood lymphocytes and it is dose-dependent.2. Cdkn1a and Gadd45a expression has been increased signigicantly by exposure of 0～5 Gray X-ray irradiation after 4 and 24 hours.3. CyclinD1 expression has been inecreased signigicantly by exposure of 0～5 Gray X-ray irradiation after 4 hours.4. X-ray irradiation induced ATM, Egr-1 and CyclinB1 gene expression shows no patterned changes or linear relationship with doses.5. Cdkn1a and Gadd45a genes have the potential to be the biomarkers of ionizing radiation.6. Gender has no effect on Cdkn1a, Gadd45a, ATM, Egr-1, CyclinB1 and CyclinD1 gene expression , but Gadd45a gene expression is impacted by smoking.
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