腺病毒与慢病毒感染猪原代细胞的比较研究

Comparative Study on Adenovirus and Lentivirus Infection of Porcine Primary Cells

本实验旨在探究腺病毒和慢病毒侵染猪原代细胞的最佳感染复数(MOI)和侵染时间,确定较优的侵染方式。实验选择体外分离培养猪骨骼肌卫星细胞、前体脂肪细胞和骨髓间充质干细胞后,分别用腺病毒(MOI=0、50、100、200、500、1 500)和慢病毒(MOI=0、30、50、80、100、300)感染细胞,每隔24 h在荧光显微镜下观察记录细胞中绿色荧光蛋白的表达情况。结果表明:当腺病毒MOI值为500、慢病毒MOI值为80,侵染4 d后,3种细胞荧光强度均较强且细胞形态完好。相比较而言,腺病毒能快速高效侵染猪骨骼肌卫星细胞;而对于猪前体脂肪细胞和猪骨髓间充质干细胞,2种病毒侵染速度相近,但慢病毒的侵染效率和荧光强度更高。因此,对于猪骨骼肌卫星细胞,选取腺病毒作为外源基因载体更为合适;对于猪前体脂肪细胞和猪骨髓间充质干细胞,慢病毒的侵染效果则更好。

This study was conducted to explore the optimal multiplicity of infection(MOI)value and harvest time of adenovirus and lentivirus infection porcine primary cells,in order to determine efficient methods for virus infection.Porcine skeletal muscle satellite cells,porcine preadipocytes and porcine bone mesenchymal stem cells were separately isolated and incubated in vitro.Adenovirus(MOI=0,50,100,200,500,1 500)and lentivirus(MOI=0,30,50,80,100,300)were respectively added to infect porcine primary cells,and the expression of green fluorescent protein were observed every 24 h using a fluorescent microscope.Results showed:when adenoviral MOI=500 or lentiviral MOI=80,after 4 d infection,the fluorescent intensity in three types of porcine primary cells was considerably strong and the cell forms remained intact.Comparatively,adenovirus could infect porcine skeletal muscle satellite cells more rapidly and efficiently.As for porcine preadipocytes and porcine bone mesenchymal stem cells(BMSCs),the infection speed of two types of virus was close,while the infection efficiency and the fluorescent intensity of lentivirus were both higher than adenovirus.Overall,adenovirus is more suitable as the gene transduction vector for porcine skeletal muscle satellite cells,while lentivirus is more efficient to infect porcine preadipocytes and porcine BMSCs.