钙信号在ALA-PDT诱导SW480细胞凋亡及细胞自身保护机制中的作用  

Role of Calcium Signal in Apoptosis and Protective Mechanism of Colon Cancer Cell Line SW480 in Response to 5-Aminolevulinic Acid-Photodynamic Therapy

作　　者：郑江华[1] 时德[1] 赵渝[1] 陈祖林[2] 

机构地区：[1]重庆医科大学附属第一医院血管外科/重庆市普外科学重点实验室,重庆400037 [2]第三军医大学附属新桥医院普外科,重庆400016

出　　处：《癌症》2006年第6期683-688,共6页Chinese Journal of Cancer

基　　金：国家自然科学基金资助项目(No.30271481)~~

摘　　要：背景与目的:肿瘤的光动力疗法(photodynamictherapy,PDT)是基于光敏剂选择性积聚在肿瘤组织中,肿瘤细胞接受可见光照后凋亡或坏死的一种治疗方法。细胞内游离钙作为重要的第二信使参与多种细胞功能的调节,但钙信号在PDT中的作用却不甚清楚。本研究的目的是探讨钙信号在!氨基酮戊酸-光动力学疗法(ALA-PDT)诱导SW480细胞凋亡及细胞自身保护机制中的作用。方法:将SW480细胞分为4组:空白对照组、激光照射组、ALA组和ALA-PDT组。用TUNEL法检测细胞凋亡;用激光共聚焦显微镜观测各组细胞内游离钙离子浓度的变化;放射免疫法测定各组细胞内cAMP及cGMP含量;半定量逆转录聚合酶链反应(RT-PCR)方法检测各组细胞中钙调素的基因表达;Western蛋白印迹检测各组细胞MEK和ERK1/2蛋白产物及磷酸化蛋白产物的表达。结果:TUNEL法显示ALA-PDT组的人结肠癌细胞在PDT后30min凋亡指数(apoptosisindex,AI)为(25.26±5.04)%,60minAI为(50.45±7.85)%;激光共聚焦结果为ALA-PDT组细胞内游离钙离子浓度在10min时荧光强度为100.00±19.83,而20min时达185.40±18.90(P<0.01),而空白对照组、单独激光照射组、ALA组细胞内钙离子浓度无明显变化;在监测时间内,cAMP含量在30min时为(3.215±0.245)pmol/L,显著高于其它各组及各组内不同时相的cAMP含量(P<0.001);空白对照组、激光照射组、ALA组以及ALA-PDT后30min、60min、90min组的CaM3的相对表达量分别为3.97±0.29、4.28±0.39、4.51±0.44、12.60±1.84、11.39±1.13和12.77±1.35,ALA-PDT后30～90min组的CaM3表达显著高于空白对照组、激光照射组和ALA组(P<0.001);ALA-PDT后SW480细胞的ERK通路激活。结论:钙信号对ALA-PDT诱导SW480细胞凋亡起着重要作用,同时也诱导了细胞自身的保护机制——ERK通路激活。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. METHODS：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. RESULTS： 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. CONCLUSIONS： Calcium signal plays an important role in ALA-PDT-induced apoptosis of SW480 cells, and can induce p

关 键 词：肿瘤 ALA PDT SW480细胞 钙信号 凋亡 

分 类 号：R730.5[医药卫生—肿瘤]