ROCK-I蛋白表达或活性的改变对人卵巢癌细胞运动表型的影响  

Effect of change of the expression or activity of ROCK-I on the motile phenotype of ovarian tumor cell.

作　　者：韩志强[1] 洪振亚[2] 陈彩虹[1] 孙立石[2] 卢运萍[1] 周剑峰[2] 马丁[1] 

机构地区：[1]华中科技大同济医学院附属同济医院妇产科,武汉430030 [2]华中科技大同济医学院附属同济医院血液科,武汉430030

出　　处：《现代妇产科进展》2007年第8期561-564,F0003,共5页Progress in Obstetrics and Gynecology

基　　金：国家自然科学基金资助项目(No:30371657);国家重点基础研究发展规划项目(No:2002CB513100)

摘　　要：目的:探讨ROCK-I蛋白表达或活性改变对人卵巢癌细胞运动表型的影响。方法:将ROCK-I的反义寡核苷酸(ASODN)或其显性激活突变体p160Δ3用脂质体介导,转染人卵巢癌细胞系CAOV-3细胞,用RT-PCR与W estern blot印迹法检测转染前后ROCK-I和p160Δ3 mRNA和蛋白的表达;rhodam ine-phalloid in染色显示ROCK-I ASODN和p160Δ3对细胞骨架的影响;Transwell小室和划痕实验观察ROCK-I表达降低及其活性提高后卵巢癌细胞系CAOV-3运动能力的改变。结果:转染ROCK-I ASODN后,CAOV-3细胞内ROCK-I蛋白的表达明显减少,最大抑制率可达49%;转染的p160Δ3在细胞内获得有效表达。转染ROCK-I ASODN后CAOV-3细胞伪足消失,肌动蛋白纤维减少且变得无序,而转染p160Δ3后细胞伪足增多变长,肌动蛋白纤维亦明显增多,且沿细胞长轴分布。伤口愈合实验显示,ROCK-I ASODN明显抑制了CAOV-3细胞的迁移,p160Δ3则显著增强了其迁移能力;转染10μmol/L或20μmol/L ROCK-I ASODN后细胞的随机运动能力分别为其对照组的(72.0±1.3)%和(55.9±2.5)%,定向运动能力分别为其对照组的(72.5±3.4)%和(54.5±1.9)%;转染p160Δ3后,与对照组相比细胞的随机运动能力提高(38.7±1.2)%,定向运动能力提高(40.2±2.6)%。结论:ROCK-I蛋白表达或活性改变与人卵巢癌细胞CAOV-3的运动能力密切相关,ROCK-I可能成为治疗卵巢癌肿瘤细胞转移的新靶点。Objective：To investigate the possible role of ROCK-Ⅰ in ovarian cancer cells locomotivity. Methods： The antisense oligodeoxynucleotide （ASODN） against ROCK-Ⅰ or its dominant-active mutant was transfected into ovarian cancer cell line CAOV-3 cells mediated by Lipofectamine 2000. The mRNA and protein expressions of ROCK-Ⅰ and p160△3 were detected by RT-PCR and Western-blot assay. To study how the changes of expression and activity of ROCK-Ⅰ ASODN and p160△3 influence the reorganization of cytoskeleton, the transfected CAOV-3 cells were stained for F-actin with rhodamine-phalloidin. Transwell chamber and woundclosure assay were used to assess the effect of ROCK-Ⅰ ASODN and p160△3 on the locomotivity of the cells. Results：The expression levels of ROCK-Ⅰ mRNA and protein in the cells were decreased significantly after transfection with 10μmol/L or 20μmol/L ROCK-Ⅰ ASODN. After transfection, p160△3 was transcripted and translated effectively in CAOV-3 cells. The cells transfected with ROCK-Ⅰ ASODN had shrunken cell shapes without the visible stress fibers. While the transfected cells with p160△3 showed pointed pseudopodia or polygonal cell shapes with pointed edges, in which the brightly and longitudinally actin bundles were formed. Woundclosure assay showed that ROCK-Ⅰ ASODN significantly inhibited the migration of CAOV-3 cells and p160△3 obviously promoted the migration of the cells. After transfection of 10μmol/L or 20μmol/L ROCK-Ⅰ ASODN ,the random migratory activity of these cells was （72.0 ± 1.3 ）% or （55.9 ± 2.5 ） % of their control group respectively, and the cells chemotaxis activity was （72.5 ± 3.4） % or （ 54. 5 ± 1.9 ） % of their control group respectively. Compared with their control group ,the random migratory activity of the cells was enhanced （38.7 ± 1.2 ）% and the chemotaxis activity was promoted （40.2 ± 2.6）% after trasfection of p160△3. Conclusions： The change of expression or activity of ROCK-Ⅰ may play a crucial role in

关 键 词：ROCK—Ⅰ 寡核苷酸类 显性激活突变体 肿瘤转移 卵巢癌 

分 类 号：R733.33[医药卫生—肿瘤]