| BackgroundWith the closer relationship between peoples' daily life and long-term low dose radiation exposure,such as medical radiological examination popularization and high frequency flight,the health effects of low dose radiation has gradually became a focus.Reliable biomarkers play a pivotal role in determining the biological effects and mechanisms of low dose radiation.At present,the most widely used biomarkers of radiation effects are cytogenetic markers represented by chromosomal aberrations,while they are limited to reflect comprehensive biological effects.Even the extremely low irradiation dose can not cause changes of cytogenetic index.Therefore,it is of great significance to find some low dose biomarkers with a wider range of appllication and a more sensitive indication.microRNA is a class of post-transcriptional regulators widely present in eukaryotes.Ionizing radiation can affect the expression of miRNAs.Meanwhile,miRNAs can also regulate the expression of various genes related to ionizing radiation,in order to be involved in the regulation of early biological effects induced by radiation,such as DNA damage and repair,cell cycle progression,cell apoptosis,and so on.ObjectiveA large number of animal experiments and cell experiments showed that ionizing radiation could induce the expression changes of miR-16,miR-106b,miR-34a,miR-449a and let-7g.However,the expression and regulation mechanism of these miRNAs are still uncertain.In this study,we select residents from high background radiation areas and AHH-1 human B lymphocytes to conduct in vivo and in vitro experimental research.In order to figure out the early biological effects and search for the effective biomarkers of low dose radiation,the expression of these five miRNAs in both peripheral blood and lymphocytes under low dose radiation were analyzed,so does the changes of miRNAs cell cycle related target genes and cell cycle.Methods110 healthy female local residents at the age of 50 years and over were randomly selected from the high background radiation area and the control area,their age,BMI and individual cumulative dose were surveyed.The relative expression level of miR-16,miR-106b,miR-34a,miR-449a and let-7g in peripheral blood plasma of these women were quantitatively detected by real-time fluorescence quantitative PCR method.Flow cytometry was used to detect the cell cycle changes of peripheral blood lymphocytes.Moreover,the AHH-1 human B lymphocytes cultured in vitro were irradiated low dose ranged from 0.05Gy,0.075Gy,0.1 Gy to 0.2G? 60Co? rays,as well as the high dose of 2Gy.Then the relative expression level of the five miRNAs were quantitatively detected by RT-PCR.The changes of cell cycle and miRNAs cell cycle related target genes including Wee1,p21,CDK1,CyclinB1 and Cdc25A were analyzed by flow cytometry.Results1.The results of high background area research(1)The levels of miR-16 and miR-106b in plasma of high background group were down regulated compared with the control group,but the level of miR-449a was up regulated(Z=-2.968,-2.089,-2.043,P<0.05).After controlling of confounding factors such as age,the expression levels of miR-16 and miR-106b were negatively correlated with the cumulative dose of individuals(?=0.243,0.208,P<0.05).On the contrary,it is not certain that high background exposure can induce the changes miR-34a and let-7g in plasma(Z=-0.414,-1.832,P>0.05).(2)The percentage of G0/G1 cells in peripheral blood lymphocytes decreased in the high background group compared with the control group,while the percentage of S-phase increased(t=3.408,-3.412,P<0.05).2.The results of AHH-1 human B lymphocytes research(1)The expression level of miR-16,miR-106b,miR-34a,miR-449a and let-7g in AHH-1 were increased along with the dose increasing after 12h induced by low dose radiation(?2=28.191,33.069,35.652,36.963,27.408,P<0.05).And there was significant difference in the relative expression level between the dose groups.The temporal trends in these five miRNAs were similar to each other,they usually hit the lowest point at 8h after irradiation,and reached a peak at 12h or 24h.Compared with the sham-irradiated group,the expression of miR-16,miR-106b,miR-34a,miR-449a and let-7g were raised to 2.548 times,6.292times,6.408times,2.501 times and 2.301 times(t=-2.667,-3.366,-5.059,-3.827,-3.739,P<0.05)at 12h with 0.075Gy irradiation.(2)The percent of G2/M phase increased at 12h and 48h after low dose irradiation in AHH-1(F=489.559,17.152,P<0.05).Meanwhile,there were significant G2/M cell cycle arrest at 2Gy irradiated.The results showed that the G2/M phase arrest was released significantly at 48h.The proportion of G2/M cells was declined from 42.77%at 12h to 7.76%.(3)The expression of p21,CDK1,CyclinB 1 and Cdc25 A decreased at 12h after low dose irradiation compared with the sham-irradiated group(F=7.957,7.934,7.833,4.040,P<0.05).And the expression of Weel,CDK1,CyclinB1 and Cdc25A were down regulated at 24h after irradiation(F=5.425,12.241,9.577,9.584,P<0.05).Conclusion1.The expression of miR-16 and miR-106b in the peripheral blood plasma of the high background area residents declined along with the individual cumulative dose increasing.It was not change that high background exposure could induce the expression of miR-449a,miR-34a and let-7g in plasma.Low dose irradiation could induce the up-regulation of miR-16,miR-106b,miR-34a,miR-449a and let-7g in AHH-1 human B lymphocytes.2.Low dose radiation exposure could increase the proportion of S phase cells in lymphocytes of high back ground area residents and G2/M phase cells in AHH-1 lymphocytes,which point out that low dose ionizing radiation induced moderate changes of cell cycle progression mediated by DNA damage.3.The expression of cell cycle related target genes including Wee1,p21,CDK1,CyclinB1 and Cdc25A,decreased in different degrees after low dose irradiation.It was illustrated that miR-16,miR-106b,miR-34a,miR-449a,let-7g and their target genes involved in the regulation of cell cycle progression induced by low dose ionizing radiation. |
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