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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 lentiviral vector containing human microRNA-203 (miR-203) and prepare a recombinant lentivirus to infect human umbilical vein endothelial cell line HU-VEC-12 and observe its effect on cell proliferation and migration. Methods The DNA fragment of miR-203 precursor was cloned by PCR and constructed and packaged into recombinant lentivirus overexpressing miR-203. The human umbilical vein endothelial cell line (HUVEC-Lv-miR-203) stably overexpressing miR-203 was established by lentivirus infection system. Real-time quantitative PCR (qRT-PCR) was used to detect the expression of miR-203. The effect of miR-203 on the growth of HUVEC in vitro was detected by MTT assay. The effect of miR-203 on migration ability of HUVEC was detected by scratch test. Results The miR-203 recombinant lentivirus overexpression system was successfully constructed. MTT assay confirmed that miR-203 can inhibit the proliferation of HUVEC in vitro. Scratch recovery test results suggest that the migration of the cell lines was significantly inhibited. Conclusion miR-203 can inhibit the proliferation and migration of HUVEC cultured in vitro.