miR-203真核表达载体的构建和重组慢病毒的制备及其对人脐静脉内皮细胞增殖和迁移的影响  

Construction and expression of recombinant lentivirus encoding miR-203 and its inhibitory effect on proliferation and migration of human umbilical vein endothelial cells

作　　者：朱华宇[1,2] 张翔[1,2] 张晓[1,2] 张瑞[1,2] 杨安钢[1,2,3] 

机构地区：[1]第四军医大学肿瘤生物学国家重点实验室,陕西西安710032 [2]第四军医大学生物化学与分子生物学教研室,陕西西安710032 [3]第四军医大学免疫学教研室,陕西西安710032

出　　处：《细胞与分子免疫学杂志》2013年第6期589-592,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(81030045;30901771)

摘　　要：目的构建含有人源microRNA-203(miR-203)的慢病毒表达载体并制备重组慢病毒,感染人脐静脉内皮细胞系(HU-VEC-12),观察其对细胞增殖和迁移的影响。方法使用PCR方法克隆miR-203前体分子的DNA片段,构建并包装过表达miR-203的重组慢病毒。利用慢病毒感染系统建立稳定过表达miR-203的人脐静脉内皮细胞株(HUVEC-Lv-miR-203)。实时荧光定量PCR(qRT-PCR)检测外源导入miR-203的表达情况。MTT法检测miR-203对HUVEC体外生长的影响,利用划痕恢复实验检测miR-203对HUVEC迁移能力的影响。结果成功构建了过表达miR-203重组慢病毒表达系统。MTT法证实miR-203可在体外抑制HUVEC增殖,划痕恢复实验结果提示该细胞株的迁移能力受到显著抑制。结论 miR-203可以抑制体外培养的HUVEC增殖和迁移。Objective To construct a recombinant lentivirus harboring human-derived microRNA-203 （miR-203）, and investigate its biological effect on proliferation and migration of human umbilical vein endothelial cells （HUVEC-12）. Methods The precursor miR-203 DNA fragment was obtained by PCR and inserted into pLenti-6.3 lentivirus vector, and then was stab- ly transfected into HUVEC-12 cells （HUVEC-Lv-miR-203）. Overexpression of exogenous pre-miR-203 was determined using real-time quantitative PCR （qRT-PCR）. Subsequently, to investigate its effects on proliferation and migration of HUVEC-12 cells, MTT assay and wound healing assay were respectively performed. Results We successfully constructed a lentivirus- based delivery system to stably express exogenous pre-miR-203 in HUVEC-12 cells. MTT assay and wound healing assay showed that miR-203 could significantly suppress cell proliferation and migration of HUVEC-]2. Conclusion MiR-203 has the inhibitory effect on the proliferation and migration of HUVEC-12 cells.

关 键 词：miR-203 人脐静脉内皮细胞 细胞增殖 细胞迁移 

分 类 号：R392.12[医药卫生—免疫学] Q256[医药卫生—基础医学]