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目的:研究离子辐射诱导Jurkat细胞凋亡的动态趋势及其机制。方法:分别以5、10、20Gy的辐射量照射Jurkat细胞,其中一份样本在照射前加入zVAD.fmk,在照射后培养6、12、24、36和48h收获细胞,应用AnVPI双染和呈现subG1峰技术,在流式细胞仪上定量测定凋亡细胞。采用WesternBlot技术检测p53蛋白的表达。结果:细胞受照射后,凋亡细胞数随培养时间的延长和辐射量的加大而增加。在6h,只有20Gy才诱导细胞显著凋亡(P<0.005),5Gy的照射要到24h才出现明显的凋亡(P<0.05)。在48h,20Gy照射诱导的凋亡细胞数是所有时间点和剂量强度中最高的(P<0.001)。加有zVAD.fmk的细胞,尽管都是10Gy照射,比未加zVAD.fmk的细胞表现出凋亡细胞明显减少(P<0.005),但与未加zVAD.fmk、照射量是5Gy的细胞相比,凋亡细胞数仍显著增加(P<0.005)。Jurkat细胞在照射前后都没有p53表达。结论:离子辐射诱导细胞凋亡呈现出时间和剂量效应关系;capase抑制剂zVAD.fmk部分抑制离子辐射诱导细胞凋亡;离子辐射诱导细胞凋亡的机制独立于肿瘤抑制基因p53,caspase可能在其中起重要作用。
Objective: To study the dynamic trend and mechanism of Jurkat cell apoptosis induced by ionizing radiation. Methods: Jurkat cells were irradiated with 5, 10 and 20 Gy respectively. One sample was added to zVAD.fmk before irradiation and harvested at 6, 12, 24, 36 and 48 h after irradiation, double staining with AnVPI subG1 peak technique to quantify apoptotic cells on a flow cytometer. Western Blot was used to detect the expression of p53 protein. Results: After irradiation, the number of apoptotic cells increased with the prolongation of culture time and radiation dose. At 6h, only 20Gy induced significant apoptosis (P <0.005), 5Gy irradiation to 24h before significant apoptosis (P <0.05). At 48h, the number of apoptotic cells induced by 20Gy irradiation was the highest at all time points and dose intensity (P <0.001). Cells treated with zVAD.fmk showed a significant decrease in apoptotic cells (P <0.005) compared with those without zVAD.fmk, even though irradiated with 10 Gy, but the cells irradiated with 5 μg of zVAD.fmk The number of apoptotic cells was still significantly increased (P <0.005). Jurkat cells did not express p53 before and after irradiation. CONCLUSION: The time-dose-dependent apoptosis-inducing effect of ionizing radiation is shown. The capase inhibitor zVAD.fmk partially inhibits the apoptosis induced by ionizing radiation. The mechanism of ionizing radiation-induced apoptosis is independent of the tumor suppressor gene p53, in which caspase may be makes an important impact.