Zheng Jiang-Hua, Shi De, Zhao Yu, Chen Zu-Lin
Department of Vascular Surgery, The First Affiliated Hospital, Chongqing University of Medical Sciences/Key Laboratory of General Surgery of Chongqing, Chongqing, 400016, P. R. China.
Ai Zheng. 2006 Jun;25(6):683-8.
BACKGROUND & OBJECTIVE: Photodynamic therapy (PDT) for tumors is based on the tumor-selective accumulation of a photosensitizer, followed by irradiation with visible light, which induces cell death and apoptosis. As an important second messenger, free calcium is involved in the regulation of several cellular processes. However, the role of calcium signal in the cells after PDT is less clear. This study was to explore the role of calcium signal in apoptosis and protective mechanism of colon cancer cell line SW480 in response to 5-aminolevulinic acid (ALA)-PDT.
SW480 cells were divided into control group, light group, ALA group, and ALA-PDT group. Cell apoptosis was detected by TUNEL assay. The changes of intracellular Ca(2+) concentration were observed by confocal laser scanning microscopy (CLSM). Intracellular cAMP and cGMP concentrations were detected by radioimmunoassay. The expression of calmodulin in SW480 cells was detected by reverse transcription-polymerase chain reaction (RT-PCR). The expression of protein products and phosphorylated protein products of MEK and ERK1/2 was detected by Western blot.
Apoptosis indexes of SW480 cells at 30 min and 60 min after PDT were (25.26+/-5.04)% and (50.45+/-7.85)%, respectively. CLSM revealed that intracellular Ca(2+) concentration was 100.00+/-19.83 at 10 min after PDT, but 185.40+/-18.90 at 20 min after PDT (P<0.01). cAMP concentration of ALA-PDT group was (3.215+/-0.245) pmol/L at 30 min after irradiation, which was significantly higher than those of other groups (P<0.001). The relative contents of CaM gene of ALA-PDT group at 30, 60 and 90 min after PDT were significantly higher than those of control group, light group, and ALA group (12.60+/-1.84, 11.39+/-1.13, and 12.77+/-1.35 vs. 3.97+/-0.29, 4.28+/-0.39, and 4.51+/-0.44, P<0.001). ERK pathway of SW480 cells was activated after ALA-PDT.
Calcium signal plays an important role in ALA-PDT-induced apoptosis of SW480 cells, and can induce protective mechanism of SW480 cells by activating ERK pathway.
肿瘤光动力疗法（PDT）基于光敏剂在肿瘤中的选择性聚集，随后用可见光照射，诱导细胞死亡和凋亡。作为一种重要的第二信使，游离钙参与多种细胞过程的调节。然而，PDT后细胞内钙信号的作用尚不清楚。本研究旨在探讨钙信号在5-氨基酮戊酸（ALA）-PDT诱导结肠癌细胞系SW480凋亡中的作用及保护机制。
将SW480细胞分为对照组、光照组、ALA组和ALA-PDT组。采用TUNEL法检测细胞凋亡。用共聚焦激光扫描显微镜（CLSM）观察细胞内Ca(2+)浓度变化。采用放射免疫分析法检测细胞内cAMP和cGMP浓度。用逆转录-聚合酶链反应（RT-PCR）检测SW480细胞中钙调蛋白的表达。用蛋白质印迹法检测MEK和ERK1/2蛋白产物及磷酸化蛋白产物的表达。
PDT后30分钟和60分钟，SW480细胞的凋亡指数分别为(25.26±5.04)%和(50.45±7.85)%。CLSM显示，PDT后10分钟细胞内Ca(2+)浓度为100.00±19.83，但PDT后20分钟为185.40±18.90（P<0.01）。照射后30分钟，ALA-PDT组的cAMP浓度为(3.215±0.245) pmol/L，显著高于其他组（P<0.001）。PDT后30、60和90分钟，ALA-PDT组CaM基因的相对含量显著高于对照组、光照组和ALA组（12.60±1.84、11.39±1.13和12.77±1.35 vs. 3.97±-0.29、4.28±0.39和4.51±0.44，P<0.001）。ALA-PDT后SW480细胞的ERK通路被激活。
钙信号在ALA-PDT诱导SW480细胞凋亡中起重要作用，并可通过激活ERK通路诱导SW480细胞的保护机制。
