Studies Of The Clustered DNA Damage Induced By Different LET Radiation And The Protection

Objective: In order to detect the DNA damages induced by different LET ionizing radiation, a molecular model was established and employed based on the conformation changes of plasmid DNA. Specific DNA damage recognizing and excising enzyme probe was used to further detect the clustered DNA damage of plasmid DNA induced byγ-rays, proton and 7Li heavy ion. The radioprotective effect of the VND3207 drug was analyzed on the DNA damages induced by different LET ionizing radiation. At last, a method was established and employed to analyze the clustered DNA damage induced byγ-irradiation in the cellular level.Methods: the irradiated plasmid DNA was digested with the E Coli for mamidopyrimidine-DNA glycosylase (Fpg) and AP endonuclease III (Endo III) which convert the damaged bases and AP sites into single-strand breaks (SSB), and then subjected to agarose gel electrophoresis. The clustered DNA damage was analyzed based on the conformation changes of plasmid DNA induced byγ-rays, proton and 7Li heavy ion to study the damages and the clustered DNA damages. In order to detect the DNA damage and the radio-protection of vanillin derivative VND3207,the irradiated liquid plasmid DNA in liquid byγ-rays, proton and 7Li heavy ion was anlyzed with the conformation changes of plasmid DNA by agarose gel electrophoresis. The genomic DNA was purified from the irradiated cells and separated under the pulsed-field gel electrophoresis after enzymes digestion, and then the clustered damage levels of the special length of DNA fragment were assessed by southern blot hybridization.Results: (1) A molecular model has been successfully established to detect the clustered DNA damage of plasmid DNA. (2)The results indicated that DNA single-strand breaks (SSB), a certain level of clustered bases lesions and a tiny of double-strand breaks (DSB) were induced by the observed 50 - 200 Gy ofγ-rays. The DNA breakage and clustered bases lesions induced by high LET proton and 7Li heavy ion were much higher as compared with that induced byγ-rays. Our data also showed that an obvious clustered DNA damage was induced by proton even at 10 Gy dose, and the effectiveness of DNA damage induction by proton was higher than 7Li heavy ion. (3) The results indicated that the protection of vanillin derivative VND3207 can effectually reduce the damages induced by the different LET ionizing radiation. Compared with the control group without drug protection, the open loop conformation of plasmid DNA of the group with VND3207 treatment was significantly reduced (p<0.01). The radioprotective effect increases along with the increasing of drug concentration. The protection of 200μmol/L VND3207 reduced the yield of 200 Gyγirradiation-induced DNA damage by 40% as contrasted with the control group. Our data indicated that VND3207 has the protective effect on the induction of DNA damage induced by irradiation from low LETγ-rays to high LET of proton and 7Li heavy ion, especially with more efficient on DNA damage induced byγ-rays and 7Li heavy ion. (4) With the method of pulsed-field gel electrophoresis (PFGE) plus southern blot hybridization, we have observed a high incidence of DSB, SSB and AP sites on the cellular genomic DNA in the cells irradiation withγ-rays.