Zhang Baoqin, Miao Lixia, Zhang Zhenxi, Xiao Mi, Chen Meilan
The Key laboratory of Biomedical Information Engineering of Ministry of Education, Pediatrics Department, First Hospital of Medical College Xi'an Jiaotong University, Xi'an 710061, China.
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi. 2005 Jun;22(3):525-9.
This experiment was designed to explore the pattern of K562 and HL60 leukemia cells death, the effects on their cell cycle and the cytoplasmic free calcium concentration ([Ca2+]i) induced by 5-aminolaevulinic acid-based photodynamic therapy (ALA-PDT). Under the transmission electron microscope (TEM), two kinds of leukemia cells' ultrastructure were observed. Flow cytometry combined with Annexin V-FITC/PI labeling was used to detect the pattern of K562 and HL60 cells' death induced by ALA-PDT. Flow cytometry combined with PI labeling was used to analyze the change in the cell cycle induced by ALA-PDT, and confocal laser scanning microscopy (CLSM) combining with calcium fluorescence probe was used to detect the change in the cytoplasmic free calcium concentration ([Ca2+]i). Immediately after irradiation, many typical apoptotic bodies were seen in the cells treated. Most of the cells treated were necrotic at 24 hours following irradiation. Flow cytometry analysis suggested that the main patterns of the cells' death were apoptosis immediately after irradiation and necrosis post-apoptosis at 24 hours post irradiation. Immediately and 24 hours after irradiation, the proportion of S phase of K562 was 57. 67% +/- 1.13% and 84.77% +/- 6.20% respectively, and the proportion of S phase of HL60 was 74.60% +/- 7.27% and 84.60% 1.74% respectively. Both [Ca+]i of the treated K562 and HL60 were increased obviously. In the best experiment condition, the initial pattern of the K562 and HL60 leukemia cells' death induced by PDT was apoptosis and the main pattern was necrosis post apoptosis. The two kinds of cells were arrested at S phase by ALA-PDT. During the death of the leukemia cells, the increase in intracellular free calcium concentration could be responsible for the ALA photodynamically induced damage to K562 and HL60 cells.
本实验旨在探讨基于5-氨基乙酰丙酸的光动力疗法（ALA-PDT）诱导K562和HL60白血病细胞死亡的模式、对其细胞周期的影响以及细胞质游离钙浓度（[Ca2+]i）的变化。在透射电子显微镜（TEM）下，观察两种白血病细胞的超微结构。采用流式细胞术结合Annexin V-FITC/PI标记检测ALA-PDT诱导的K562和HL60细胞死亡模式。采用流式细胞术结合PI标记分析ALA-PDT诱导的细胞周期变化，并用共聚焦激光扫描显微镜（CLSM）结合钙荧光探针检测细胞质游离钙浓度（[Ca2+]i）的变化。照射后立即观察到，处理后的细胞中出现许多典型的凋亡小体。照射后24小时，大多数处理后的细胞发生坏死。流式细胞术分析表明，细胞死亡的主要模式在照射后立即为凋亡，照射后24小时为凋亡后坏死。照射后立即和24小时，K562细胞S期比例分别为57.67%±1.13%和84.77%±6.20%，HL60细胞S期比例分别为74.60%±7.27%和84.60%±1.74%。处理后的K562和HL60细胞的[Ca+]i均明显升高。在最佳实验条件下，PDT诱导的K562和HL60白血病细胞死亡的初始模式为凋亡，主要模式为凋亡后坏死。ALA-PDT使两种细胞停滞于S期。在白血病细胞死亡过程中，细胞内游离钙浓度的升高可能是ALA光动力诱导K562和HL60细胞损伤的原因。
