Mechanistic Study Of Radiosensitization By Arsenic Trioxide On Human Hela Cell Line Of Cervical Cancer

IntroductionCervical carcinoma is a most common gynecological malignancy. Radiation therapy can cure or relieve it among part of the patients, while the prognoses a-mong other patients treated solely by the therapy are not good. A new type of treatment named cervical carcinoma sensitivity - increasing radiotherapy attracts the great attention of clinical researchers. As₂O₃ is also called arsenic or arseni-ous acid. Its a kind of poison and has long been considered as a kind of carcinogenic drug. In 1970s, its firstly used to cureAPL and had good curative effect, therefore, its approved by FDA to cure the disease. There are many fundamental research reports about its harm to cells. As₂O₃ induces the polarization of the malignant cells to change their cycles, which leads to the apoptosis of them and inhibit the creation of tumor vein to realize antineoplastic functions. The above changes made by As₂O₃ may increase the radiation sensitivity of the tumor. Therefore, this experiment observes whether As₂O₃ can kill the cervical cancer cells by way of increasing radiation. What's more, elementary research of increasing the radiation sensitivity mechanism is made to provide the experimental reference to the treatment of cervical carcinoma using As₂O₃ combined with radiation therapy.Materials and methods1. Materials sourcesAs2O3 injection liquid is purchased from Har bin Yi Da pharmaceutical factory.Cervical cancer Hela cells are purchased from the research institute of China Medical University.2. MethodsMTT assay is used to measure the surviving rate of the Cervical carcinoma Hela cells. Then calculate the half inhibitive dosage( ID50) value. Colon forming assay is carried out to take count of survival fraction. Cell survival curve is analyzed using Multi - target single - hit model and then SER is calculated. Flow cytometry is performed to access the influnce of As2O3 on cell cycle distribution and apoptosis.3. Statistical analyses: Use the statistical software of SPSS11.5 and Graph-Pad prism to process the data.Result1. The inhibition of As2O3 acing on Hela cells of cervical carcinoma increases with the increase of concentrations. ID50;19. 92umol/L.2. The average deadly dosage (Do) of Hela cells treated by medication decreases (1. 58Gy ^2. 11 Gy ) , the Dq value is less than that in the sole irradiation group (0.27 Gy^0.64 Gy) ,and the surviving curve Dq becomes smaller. Sensitization enhancement ratio (SER) is 1.34.3. The cells proportion in G2/M period increases gradually after treated by As2O3, the proportion in Gq/Gj and S period decreases gradually, and the apoptosis increases.DiscussionAs2O3 is called arsenic or arsenious acid. Its a kind of poison and has long been considered as a kind of carcinogenic drug. In 1970"s, it's firstly used to cure APL. The present clinical indication is not limited to cure the leukemia, and the clinical application for carcinoma is being carried out overseas. The inhibition of As2O3 acting on tumor is as follows, 1. Adjust the tumor cell cycle, prevent the cells in G2/M period, and make cells more sensitive to radiationtherapyo 2. Induces the polarization of the tumur cells. 3. Induce the apoptosis of the tumor cells. Inhibit the occurring and transferring of the tumor mlti - target. 4. Inhibit the occurring of new tumor vein.Measure the cells surviving rate with MTT measurement. The result is that the As2 03 influence on cervical carcinoma Hela cells is in compliance with the dose, which means the inhibition ratio of the cells increases with the concentration, ID50:19. 92umol/L. In the following sensitivity -enhancing experiment, the concentration of 4umol/L is adopted, which is no harm to the cells.The classical cells surviving curve is educed with multi - target single -hit, and the values of Do and Dq are calculated. It can be seen from the surviving curve data that the surviving curve Dq of the cells irradiated with 4umol/L As2O3 becomes small, the value of Dq is less than that in the sole irradiation group (0. 27 Gy^O. 64 Gy) , the value of Do is less than that in the sole irradiation group ( 1. 58GyN2. 11 Gy) , and SER is 1. 34. The value of Dq denotes the cells repair ability to the sublethal damage of radiation, so the above results indicate that As2O3with the function of radiation sensibility - enhancing can inhibit the repair ability of Hela cells and decrease the cells surviving rate after irradiating.The cells radiation sensibility is related not only to the inherent DNA rehabilitating mechanism, but also to the cell cycle. It is the most sensitive period of M and G2 period in cell cycle. In the late S period, the cells resist the radiation very much. The cells radiation sensitivity is in medium in Gj period and early S period. The analyzing result by the flow cytometry in this experiment shows that the ratio of Hela cells treated by As2O3 in G2/M period decreases gradually with the increase of medicament concentration. The above results indicate that the induced cell cycle rearrangement is probably a radiation sensibility - enhancing mechanism of As2O3.The analyzing results by the flow cytometry in this experiment shows that the apoptosis of Hela cells treated by As2O3 becomes obvious peak before Gj period. The number of the apoptosis cells increase with the increase of concentration. The research result is that the above phenomenon is related with the expression levels of p53 gene, bax gene and other related genes. The tumor cellstreated by As2O3 not only apoptosis, but also get into a high expression status, which makes it easier to induce the cells with radiation to their apoptosis and shows higher radiation sensitivity.This experiment shows that As2O3 has the obvious radiation sensibility - enhancing function on cervical carcinoma Hela cells. It is presumed that As2O3can probably change the radiation sensitivity through inhibiting the Hela cell repair of sublethal damage, changing its cycle and finally inducing its apoptosis .Conclusion1. As2O3 has obvious radiation sensibility - increasing function on Helacells.2. As2O3 can change the radiation sensibility of the cells probably by way of inhibiting Hela cells repairing, changing the cell cycle and induced cells apoptosis.