Study Of Adenovirus E1A Gene Expression On Cell's Radiosensitivity

Objective: To evaluate the effect of E1A gene expression on cell's radiosensitivity. Methods: pcDNAIII vector containing E1A gene was transfected into human lung cancer cells (Anip-973), then the transfected cells were selected by G418, the positive clone was identified by PCR, RT-PCR and immunocell-chemical stain. Single dose irradiation with 6MV-X rays of 0Gy, 1Gy, 2Gy, 3Gy, 5Gy, 7Gy, and 10Gy, respectively were given to the Anip-973 cells, Anip-973 cells transfected with vacancy vector (Anip-973-vect), and the Anip-973 cells transfected with E1A gene (Anip-973-E1A) to make survival curves. Furthermore, survival ratio assays of Anip-973, Anip-973-vect, Anip-973-E1A were made with MTT method after 5Gy single fraction irradiation, cell cycle distribution and apoptosis were analysised by FCM. Results: E1A gene has integrated into positive transfected cells and expressed stably confirmed by PCR, RT-PCR and immunocell-chemical stain. Cell survival-curves showed: D₀ values of Anip-973 cells, Anip-973-vect cells and Anip-973-E1A cells were 1.14, 1.13, 1.02 respectively and Dq values were 1.53, 1.44, 0.8 respectively. The survival curves of Anip-973 cells and Anip-973-vect cells did not show significant difference, but the survival curve of Anip-973-E1A cells showed narrow shoulder region, steep slope, a radiosensitive ratio of 1.12 was obtained. The lowest survival rates after 5 Gy irradiation were at 14 hour, survival rates of Anip-973 cells and Anip-973-E1A cells were 69.5% and 41.5% respectively (p< 0.05). The results of FCM showed there were G₂ stage delays in Anip-973 cells and Anip-973-vect cells after 5Gy irradiation, while there was neither G, stage nor G₂ stage delay in Anip-973-E1A cells. Only the Anip-973-E1A cell occured obvious apoptosis at 30 hour after 5Gy irradiation. Conclusions: E1A gene increases interphase death and proliferative death and thus enhances radiosensitvity of Anip-973 cells by decreasing repaire of sublethal damage, abrogating the dalay of G₂ stage and increasing apoptosis.