Non-thermal Plasma Enhances Radiosensitization In Tumor Cells By Arresting G2/M Phase And Affecting Related Cell Cycle Protein

Objective:Non-thermal plasma(NTP)is a kind of mixed gas in the fourth form.It includes ions,electrons,neutrons and free radicals.We have done lots of experiments to confirm that non-thermal plasma can selectively kill cancer cells,while protecting normal ones both in vivo and in vitro.And convincing evidence has been accumulated that low dose of NTP shows an obvious effect of radiosensitization in a variety of tumor cells.However,the mechanism of this effect on cancer cells is still not clear,and further study is needed.Therefore,this paper intends to explore the mechanism of X-ray irradiation and adjuvant non-thermal plasma to affect related G2/M cell cycle arrest proteinMethods:1、NTP device and X-ray irradiation apparatus parameters were set by the previous experimental results.And tumor cells were divided into four groups:Control group(Cont.),Non-thermal plasma alone group(NTP),X-ray irradiation alone group(IR),Non-thermal plasma joint X-ray irradiation group(NTP+IR).2、The expression of γ-H2 AX in Hep-G2 and HeLa cells treating with non-thermal plasma joint X-ray irradiation was investigated using immunofluorescence staining technology.3、The cell cycle distribution of Hep-G2 and HeLa cells after combined treatment was measured using flow cytometry technique.4、The effect of related cell cycle protein in Hep-G2 and HeLa cells after combined treatment was detected using Western Blot technology.5、A successful ATR Ri Hep-G2 cell model was built using plasmid amplification andcell transient transfection technology;the effect of related cell cycle protein in ATR Ri Hep-G2 cells after combined treatment was detected using Western Blot technology.Results:1、The expression of γ-H2 AX in Hep-G2 and HeLa cells reached a peak both at 6h.The expression in combined treatment was significantly increased at 12 h and 24 h,compared with that of the X-ray alone group.2、Hep-G2 and HeLa cells had a slight effect on G2/M phase in NTP group.There was a G2/M phase cell cycle arrest phenomenon in IR group,while the tendency was more remarkable in NTP+IR group.3、The cell cycle arrest was disappeared in ATR Ri Hep-G2 cells with NTP,while the G2/M cycle arrest still appeared in ATR Ri Hep-G2 cells with X-ray irradiation.Conclusion:1、The DNA bear much more damage with combined treatment,and the duration of damage was prolonged.2、The combined treatment can induce more G2/M phase tumor cells,and the target is regulated by Cdk1-Cyclin B1 complex.3、NTP caused G2/M phase block effect is related to the ATR gene.