水通道蛋白4重组质粒的构建及其在体外对低糖反应的观察  

Construction of p EGFP-C2-AQP4-M23 plasmid and the effect of AQP4 on astrocytes in hypoglycemia

作　　者：俞晓燕[1] 邓江山[1] 赵玉武[1] 

机构地区：[1]上海交通大学附属第六人民医院,200233

出　　处：《实用医学杂志》2015年第5期700-704,共5页The Journal of Practical Medicine

基　　金：国家自然科学基金资助项目(编号:31271125)

摘　　要：目的:构建大鼠水通道蛋白4(aquaporin4,AQP4)重组质粒,检测融合蛋白在CTX-TNA2星形胶质细胞上的表达,同时观察低浓度葡萄糖条件下AQP4对细胞的影响。方法:通过RT-PCR方法从大鼠脑部获取AQP4基因,插入p EGFP-C2质粒中得到p EGFP-C2-AQP4-M23重组质粒,同时转染CTX-TNA2细胞后用不同浓度葡萄糖处理,通过激光共聚焦显微镜检测AQP4在细胞中的表达及低糖条件下对细胞的影响。结果:PCR和酶切鉴定结果显示插入的AQP4基因大小正确,测序结果证实成功构建了p EGFP-C2-AQP4-M23重组质粒;在激光共聚焦显微镜下观察到AQP4主要位于CTX-TNA2细胞膜上;低糖时转染AQP4的星形胶质细胞平均表面积明显高于对照组。结论:本研究成功构建了AQP4重组质粒并在CTX-TNA2细胞上成功表达,同时证实低糖时AQP4参与了星形胶质细胞水肿的形成。Objective To construct pEGFP-C2-AQP4-M23 plasmid and to detect its expression in CTX-TNA2 cells in order to study the effect of AQP4 on astrocytes in hypoglycemia. Methods AQP4 gene obtained from SD rats by RT-PCR was cloned into pEGFP-C2 plasmid. The recombinant plasmid was transfected into CTX-TNA2 cells, then treated with low glucose. The expression and effect of AQP4 on CTX-TNA2 cells were examed with confocol. Results PCR and enzyme digestion showed AQP4 size was right; confocol demonstrated that AQP4 was expressed on the membrane of CTX-TNA2 cells, and the surfacial area of CTX-TNA2 cells with AQP4was larger than those without AQP4 in hypoglycemia. Conclusion pEGFP-C2-AQP4-M23 plasmid was successfully constructed and expressed in the CTX-TNA2 cell membrane and AQP4 plays an important role in cell edema in hypoglycemia.

关 键 词：水通道蛋白质4 绿色荧光蛋白质类 转染 细胞水肿 

分 类 号：R3411[医药卫生—基础医学]