Biological Effects Of Nonthermal Plasma Combined With X-ray Irradiation On Hep-G2 Liver Tumor Cells

Part 1:Synergistic effect of Nonthermal Plasma combined with X-ray on Hep-G2CellsPurpose:To observe the effect of Nonthermal Plasma on Hep-G2 cells at different time.Methods:The inhibition rate of Hep-G2 cells was detected by CCK-8 kit at 0s,5s,15s,20s,40s,80s,150s and 300s,and the inhibition curve was used to find the Nonthermal Plasma non-toxic dose.Results:(1)When the low temperature plasma irradiation time is 20s,the cell inhibition rate is close to 0,which can be regarded as non-toxic dose point.(2)The apoptotic ratio of Hep-G2 cells in combination therapy group was higher than that in X-ray irradiated group(P<0.01,=0.02,0.01),and 24h and 72h after exposure to24h,48h and 72h Compared with the time point,the difference of apoptotic rate was significant(P=0.005).Conclusion:Nonthermal Plasma combined with 2Gy X-ray irradiation has synergistic effect on Hep-G2 cells.Part 2:Biological effects of nonthermal plasma combined with X-ray irradiation on human hepatocellular carcinoma in nude micePurpose:To investigate the biological effects of nonthermal plasma(NTP)combined with X-ray irradiation on human hepatocellular carcinoma in nude mice.Methods:By subcutaneous Hep-G2 cell inoculation,tumor model of human hepatocellular carcinoma Hep-G2 in nude mice were established,and these mice were divided into control group,NTP treatment group,X-ray irradiation group,combined treatment group.The changes of tumor microenvironment were observed using hematoxylin-eosin(HE)staining.The terminal deoxynucleotidyl transferase-mediated nick end labeling(TUNEL)was used to determine the level of neuronal apoptosis in tumor tissues.The expression of metal matrix proteinase-2(MMP-2)was detected by immunohistochemical assay.High power transmission electron microscope was used to observe cell structure changes.Results:(1)The final tumor volumes of control group,NTP treatment group,X-ray irradiation group,combined treatment group were(543.96±108.45),(436.54±65.49),(351.66±56.68),(281.97±35.60)mm~3,with a statistically significant difference(F=9.63,P=0.01),and the difference between X-ray radiation group and the combination treatment group was significant(P=0.05).(2)HE staining showed that the combined treatment group compared with the other groups,there was a larger area of necrosis.(3)TUNEL showed that the apoptotic indexes were(0.95±0.13)%,(5.82±0.26)%,(7.53±0.43)%,(11.07±0.35)%respectively,with a statistically significant difference(F=547.76,P<0.01),and the difference between X-ray radiation group and the combination treatment group was statistically significant(P<0.01).(4)The immune scores of each group were 12,9,9,2.(5)High power electron microscopic observation showed that there were more apoptotic bodies in the combined treatment group than those in the other groups,accompanied by mitochondrial edema.Conclusion:NTP and X-ray irradiation therapy in the treatment of human hepatocellular carcinoma in nude mice model has a synergistic effect,which probably provides a new treatment for malignant tumors.