摘要
目的用siRNA沉默ClC-3氯通道,探讨ClC-3氯通道在鼻咽癌CNE-2Z细胞调节性容积回缩(regulatory volumedecrease,RVD)中的作用。方法使用终浓度100 nmol/L的ClC-3 siRNA转染鼻咽癌CNE-2Z细胞,流式细胞术检测siRNA的转染率,Western blotting检测ClC-3蛋白表达,活细胞图像系统分析对照组和ClC-3 siRNA组细胞在低渗(160mOsm/L)刺激时的容积调节能力。结果 ClC-3 siRNA转染率为(63.8±3.8)%(n=3,P<0.01),与对照组相比,ClC-3氯通道蛋白的表达减少(60.9±4.0)%(n=3,P<0.01)。对照组细胞的RVD为(42.6±2.8)%(n=20),ClC-3 siRNA组细胞在低渗液中的最大容积与对照组没有显著性差异,但RVD仅为(10.5±4.8)%,RVD抑制率达75.4%(P<0.01)。结论 ClC-3 siRNA成功转染鼻咽癌CNE-2Z细胞,特异性抑制ClC-3蛋白表达,显著降低细胞RVD能力,提示ClC-3氯通道在鼻咽癌细胞调节性容积回缩中起重要作用。
Objective To investigate the role of ClC-3 chloride channels in regulatory volume decrease（RVD） of nasopharyngeal carcinoma（NPC） CNE-2Z cells.？Methods ClC-3 siRNA was transfected into CNE-2Z cells in the presence of the transfection reagent HiPerFect ReagentTM.The transfection efficiency of ClC-3 siRNA was detected by flow cytometry.The expression of ClC-3 protein was detected by Western blotting,and the changes of cell volume in 160 mOsmol/L hypotonic solution were determined by image analysis.？Results The transfection efficiency of ClC-3 siRNA was（63.8±3.8）%（n=3,P0.01）,and compared with the control group,ClC-3 siRNA transfection resulted in a reduction of ClC-3 expression by（60.9±4.0）%（n=3,P0.01）.The hypotonic challege（160 mOsmol/L） caused cell swelling and induced RVD.In the control group,hypotonic solution bath for 35 min resulted in a RVD of（42.6±2.8）%（n=20）,which was significantly decreased to（10.5±4.8）%（n=16） in ClC-3 siRNA-transfected cells,demonstrating a reduction of RVD capacity by 75.4%（P0.01）.？Conclusion The capacity of RVD is significantly reduced in CNE-2Z cells by ClC-3 chloride channel protein knock-down via ClC-3 siRNA transfection,indicating an important role of ClC-3 chloride channels in the RVD of CNE-2Z cells.
出处
《南方医科大学学报》
CAS
CSCD
北大核心
2011年第2期216-220,共5页
Journal of Southern Medical University
基金
国家自然科学基金(30771106
30870567
30871267)~~
关键词
ClC-3氯通道
SIRNA
鼻咽癌细胞
细胞容积
ClC-3 chloride channels
small interfering RNA
nasopharyngeal carcinoma cells
cell volume
