超声激活血卟啉抑制肝癌细胞抗失巢凋亡能力的机制初探  

Effect of Singlet Oxygen on Inhibiting Anoikis Resistance in Hepatocarcinoma Cells Via Decreasing Expression of Osteopontin

作　　者：成涓[1] 冉海涛[1] 王志刚[1] 许川山[1] 李攀[1] 谢昭鹏[1] 张茂惠[1] 蒲世玉[1] 

机构地区：[1]重庆医科大学超声影像学研究所,重庆市400010

出　　处：《中国超声医学杂志》2008年第6期488-491,共4页Chinese Journal of Ultrasound in Medicine

基　　金：国家高技术研究发展计划(863计划No.2006AA02Z4F0);国家自然科学基金面上项目(No.30770565)

摘　　要：目的探讨血卟啉经超声激活后能否生成单线态氧、降低骨桥蛋白(osteopontin,OPN)表达,从而抑制肝癌细胞株HepG2的抗失巢凋亡能力。方法将HepG2细胞悬液随机分为5组:血卟啉组,培养液中加入血卟啉40μl孵育30 s;超声辐照组,以1.0MHz、1.5W/cm2超声辐照30 s;血卟啉与超声辐照组,同样方法予以血卟啉与超声联合处理;血卟啉、组氨酸与超声辐照组,加入活性氧捕获剂-组氨酸后进行血卟啉与超声联合处理;及空白对照组。各组处理后细胞悬液一部分利用1,3-二苯基异苯并呋喃(1,3-diphenylisobenzofuran,DPBF)估计单线态氧生成量,一部分在软琼脂中培养2周后观察其集落形成能力,还有一部分经悬浮培养后以Annexin V-FITC/PI双染法检测失巢凋亡率、RT-PCR检测骨桥蛋白的mRNA表达、Western blot检测OPN的蛋白表达。结果血卟啉与超声辐照组DPBF的相对消耗量即单线态氧生成量明显高于其它各组,且该组HepG2细胞在软琼脂中无细胞集落形成,与其它组相比失巢凋亡率增高(18.46%±4.08%)、OPN的表达降低。结论血卟啉经超声激活后产生的单线态氧可降低OPN表达,从而发挥其抑制HepG2细胞抗失巢凋亡的作用。Objective To investigate the mechanism of hematoporphyrin activated by ultrasound for inhibiting the anoikis resistance in Hepatocarcinoma cells. Methods The HepG2 cell suspension was randomly divided into five groups as follows： the cells in hematoporphyrin group were incubated in culture solution containing 40 microliters of hematoporphyrin for 30 s; those in ultrasound group were irradiated by 1.0 MHz, 1.5 W/cm^2ultrasound for 30 s; those in hematoporphyrin and ultrasound group were treated by both; those in histidine, hematoporphyrin and ultrasound group were treated by histidine treated with hematoporphyrin and ultrasound and others without treatment as the control group. After treatment singlet oxygen was proved by measuring the relative loss of diphenylisobenrofuran （DPBF） . A number of cells were cultured in soft agar for two weeks to observe their growth ability when losed cell adhesion. Other cells were cultured in plates covering with poly-HEMA and the anoikis was detected by Annexin V-FITC/PI double staining. The expression of osteopontin （OPN） was detected by reverse reverse transcription-polymerase chain reaction （RT-PCR） and Western blot analysis. Results In hematoporphyrin and ultrasound group, the relative loss of DPBF was increased, there was no cell colony in soft agar, the apoptosis rate was 18.46±4.08%, and the expression of OPN was decreased. Differences from other four groups were significant. Conclusions Singlet oxygen generated by hematoporphyrin and ultrasound irradiation can inhibit the anoikis resistance in HepG2 hepatocacinoma cells via decreasing the expression of OPN.

关 键 词：肝脏 肿瘤 单线态氧 失巢凋亡 1 3-二苯基异苯并呋喃 骨桥蛋白 

分 类 号：R735.7[医药卫生—肿瘤] R758.630.3[医药卫生—临床医学]