携载增强型绿色荧光蛋白基因慢病毒转染骨髓间充质干细胞示踪方法

Study on Bone Marrow Mesenchymal Stem Cells being Transfected with Lentivirus Carrying Enhanced Green Fluorescent Protein Gene

目的本实验研究旨在研究使用携带增强型绿色荧光蛋白(enhanced Green Fluorescent Protein,eGFP的慢病毒转染骨髓间充质干细胞(bone marrow mesenchymal stem cell,BMSCs)的方法,用于解决组织工程骨研究中种子细胞分化、增殖过程中的示踪问题。方法用携带eGFP的慢病毒转染第3 代BMSCs,用流式细胞仪检测转染率,获得稳定表达eGFP 的BMSCs,并通过描绘生长曲线来判定对BMSCs 增殖性能的影响。结果当MOI值为70 时,经eGFP 慢病毒转染后的BMSCs 能够得到活性较好、转染率高的eGFP-BMSCs;转染后各细胞生长曲线呈"S"形,MOI=70 与MOI=0 时细胞增殖最好,MOI=100 会对BMSCs 增殖造成影响。结论MOI=70 时携载eGFP基因慢病毒转染BMSC效果最好。

Objective The purpose of this study was to investigate the transfection of bone marrow mesenchymal stem cells with lentivirus carrying enhanced green fluorescent protein and to solve the tracer problem in the process of seed cell differentiation and proliferation in tissue engineering bone research. Methods The third-generation BMSCs were transfected with lentivirus carrying eGFP,and the transfection rate was determined by flow cytometry. BMSCs with stable expression of eGFP were obtained, and the effect on proliferation of BMSCs was determined by growth curves.Results Under MOI=70,BMSCs transfected with eGFP lentivirus showed better activity and higher transfection rate.After transfection,the growth curve of each type of cell presented an "S" shape.The proliferation of BMSCs was best at MOI=70 and MOI=0,it was affected at MOI=100.Conclusions Lentivirus carrying eGFP gene transfected BMSC had the best effect at MOI=70 in vivo.