【摘要】 探讨基于氨乙酰丙酸介导的光动力作用(ALA-PDT)对白血病细胞株HL60和K562细胞的杀伤模式及其可能机制。在ALA-PDT处理细胞的优化条件下,用透射电镜对处理细胞进行超微结构观察,选择AnnexinV-FITC/PI双标法明确ALA-PDT对两种白血病细胞的杀伤模式,PI染色流式细胞仪分析ALA-PDT后细胞周期的变化,以Fluo-3/AM为钙离子指示剂采用激光共聚焦显微镜检测细胞内钙离子浓度的变化。透射电镜观察发现PDT后即刻的细胞呈现典型的凋亡小体,24h后大部分发生坏死;双标法检测显示PDT后即刻及24h后细胞的死亡模式分别以凋亡和凋亡后坏死为主。光动力作用后即刻和作用后2h,K562细胞S期所占的比例分别为57.67%±1.13%和84.77%±6.20%,HL60细胞S期所占的比例分别为74.6%±7.27%和84.60%±1.74%;两种细胞内钙离子浓度的均明显升高。在最佳试验条件下,凋亡是ALA-PDT杀伤白血病细胞K562和HL60细胞的最初模式,而凋亡后坏死为其主要模式,ALA-PDT使白血病细胞株阻滞于S期,在白血病细胞株死亡的过程中,细胞内钙离子浓度的增加可能起着重要的作用。更多还原

【Abstract】 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 ([Ca 2+]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 ([Ca 2+]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 2+]_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.更多还原