Effects Of Radiosensitization Of Reverse Transcriptase Inhibitor On Human Malignant Glioma Cells U251

Objects To study radiosensitization of reverse transcriptase inhibitor (3'-azido-3'- deoxythmidine, AZT), we investigate the effects of AZT combined with y-irradiation on telomerase activity, elomere length, DNA single-strand breaks and DNA double-strand breaks induced by irradiation in human malignant glioma cells U251.Methods Exponentially growing U251 cells were devided into four groups: A: control, B: to be treated with irradiation(2Gy), C: to be treated with 0.8mm/L AZT for 24h, D: to be treated with AZT and irradiation both. The telomerase activity was determined by telomeric repeat amplification protocol (TRAP-PCR-ELISA); telomere length was determined by Southern Blot; DNA damage after irradiation was determined by alkaline or neutral single cell gel electrophoresis assay(comet assay). The changes in radiosensitivity was quantified by calculating the sensitization enhancement ratio (SER) through survivle curves. SERSF2 was defined as the rate of survivle fractions at 2Gy(SF2) for irradiation and the SFi for AZT combined with irradiation.Results AZT resulted in an decrease in telomerase activity in both radiated and unradiated cells(1.121?.161, 1.019?.116 vs 1.567?.037,P<0.05). Telomere lengeth decreasing was investigated in radiated cells but not in unradiated cell. There was a statistically significant difference between group D and group B in telomere length at 15min,30min,1h after irradiation(P<0.05). Immediately after irradiation, the type comet image was observed in radiated cells but not in unradiated cells, as well. There was no significantly differences in the level of initial SSB and initial DSB between group B and group D. While the rates of repair of SSB and DSB were slower in group D than that in group B(P<0.05). The irradiation induced DNA SSB and DSB were rather efficiently repaired in group B within 2h, while there were still about 50% residual SSB and 40% residual DSB in group D at that time(P<0.05). Through survivle curves we obtained DQ>Dq, SF2 of U251 cells: 1.803, 2.5, 0.752, respectively. SERD0, SERDq SERSF2 for AZT were 1.294, 1.25, 1.365 respectively. Correlation assay had showed that telomere lenth, repair efficiency of SSB and DSB was highly correlated with telomerase activity (r>0.7).Conclusions AZT can increase the radiosensitivity of U251. The mechanism of radiosensitization may be related to the inhibition of telomerase activity, which willeffect the subsequent repair of irradiation induced telomere shorting and DNA damage.