辐 射 研 究 与 辐 射 工 艺 学 报 V ol.23, No.2 94 J. Radiat. Res. Radiat. Process. April 2005 ——————————————
Effects of nitroprusside sodium to prevent cell cycle arrest induced in
A172 human glioma cells by X-ray irradiation
QIU Rong LI Wenjian HAO Jifang LIU Bing ZHANG Hong
(Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000)
(Radiotherapy Department, General Hospital of Lanzhou Military Area, Lanzhou, 730050)
Nitric oxide (NO) has been implicated both in regression and progression of tumors due to its production by both tumor cells and infiltrating leukocytes. Ionizing radiation causes the regression of tumors, and can augment the production of NO by macrophages in vitro. The authors investigated the effect of X-ray irradiation on cell cy-cle of A172 human glioma cells pretreated with nitroprusside sodium (NPS), the NO donor.
Human glioma A172 cells were maintained in RPMI-1640 medium supplemented with 10% heat-inactive fetal calf serum in a 5% CO 2 incubator at 37℃. The experiment was performed in exponentially growing cultures. The cells in cell culture flasks treated with or without 1mM NPS for 4h. were irradiated to a dose of 4Gy X-rays using 8MeV CL 2100C electron linear accelerator (Varian, USA) in General Hospital of Lanzhou Military Area. After the irradiation, the cells were collected by trypsinization, washed with phosphate buffer solution, fixed in 70% ethanol, treated with RNase and the nuclei were stained with propidium iodide. The DNA content was evaluated using Becton Dickinson FACSCalibur and analyzed with UMCM cylchred cell cycle analyzer software.
The cell culture medium was replaced after irradiation and cells were harvested 24h later, labeled with propidium iodide and analyzed by flow cytometry. After the 4Gy X-ray irradiation, the cell cycle distribution of A172 glioma cells changed markedly, with increased percentage of G 2/M cells and decreased of G 0/G 1 and S cells compared to control cells(Fig.1a, 1b). The cells pretreated with 1mM NPS 4h displayed similar cell cycle distri-butions to those of the control (Fig.1c). But X-ray irradiated cells preincubated with 1mM NPS showed different cell cycle distributions from those of unpretreated cells (Fig.1d).
Fig.1 Cell cycle distributions of human glioma cells
b. 4Gy X-ray irradiation,
c. 1mM NPS 4h,
d. 1mM NPS 4h+4Gy X-ray irradiation
It was reported that NO or NO release agents were as effective as oxygen to radiosensitize hypoxic cells in vitro. It was also revealed that NO involved in the resistance to ionizing radiation by by-stander effects. The re-sults in the present research demonstrated that NO donor inhibit A172 glioma cell undergo cell cycle arrest fol-lowing X-ray irradiation. Several studies have shown that p53 protein accumulates in the nuclei of cells exposed to ionizing radiation. Whether impairing p53 function involved in prevention of NO to cell cycle arrest induced by X-ray remained further investigation.
KEYWORDS Nitric oxide, Glioma, Radiation, Cell cycle