摘要
目的建立稳定高表达和靶向干扰谷氨酰胺合成酶(GS)细胞株观察GS对C6胶质瘤细胞增殖、迁移和侵袭能力的影响。方法将已构建的GS-pEGFP-N3、GS-shRNA真核表达载体和空载体pEGFP-N3通过脂质体介导分别转染C6胶质瘤细胞株,G418持续筛选并用Western blot鉴定,MTS法和克隆形成实验观察GS对C6胶质瘤细胞增殖的影响,用MTS黏附实验、划痕损伤实验和transwell侵袭实验检测GS对细胞黏附、迁移和侵袭能力的影响。结果 Western blot证实过表达细胞株有超量GS蛋白表达,而shRNA细胞株未检测到GS信号;GS过表达细胞增殖、克隆形成能力和侵袭性显著降低而黏附能力显著增强;shRNA细胞增殖和克隆形成能力无显著性变化,黏附能力显著降低,而侵袭性显著增强。结论成功建立GS稳定高表达和靶向干扰C6胶质瘤细胞株,并证实GS表达水平影响胶质瘤细胞的增殖、迁移和侵袭。
Objective To study the effects of glutamine synthetase （GS） on proliferation,migration and invasion of C6 stable cell lines which transfected with GS expression vector or shRNA vector. Methods C6 stable cell lines were generated by introducing GS shRNA,GS-EGFP-N3 or EGFP-N3 vector and selected with G418 followed by cloning. GS expression was detected by Western blot. Cell proliferation was detec- ted by MTS and colony-forming experiment. Adhesion assay, wound healing and transwell experiments was used to detect the cell adhesion,migration and invasion. Results Western blot showed that GS was over-expressed in GS stable ceils and no signal was detected in GS knocking down cells. GS over-expres- sion suppressed cell growth, clone formation and invasion, increased cell adhesion; GS knocking-down increased cell invasion,decreased cell adhesion,and had no significant impact on cell growth and clone formation. Conclusion The level of GS affects cell growth,migration and invasion of C6 glioma cells.
出处
《肿瘤防治研究》
CAS
CSCD
北大核心
2012年第10期1197-1201,共5页
Cancer Research on Prevention and Treatment
基金
国家自然科学基金资助项目(81000527)
江苏省自然科学基金资助项目(BK2010159)
上海市华东疗养院院级基金资助项目(201001)
关键词
谷氨酰胺合成酶
C6细胞株
稳定表达
增殖
迁移
侵袭
Glutamine synthetase
C6 glioma cell line
Stable expression
Proliferation
Migration
Invasion
