重组腺相关病毒对NK92细胞的转导条件优化

Optimization of transduction conditions of recombinant adeno-associated virus into NK92 cells

为提高重组腺相关病毒(recombinant adeno-associated virus,rAAV)对NK92细胞的转导效率,对转导细胞密度、培养基中IL-2浓度、rAAV血清型和用量进行优化;此外,用不同浓度的促进剂(ZnCl_(2)、氯喹、聚乙烯醇和金雀异黄酮)溶液处理细胞,以进一步提高病毒转导效率。结果表明,在细胞密度为5×10^(5)时,增强型绿色荧光蛋白(EGFP)的表达效率相对较高;当培养基中IL-2的浓度为1000 IU/mL时,NK92的细胞生长状态最适合病毒转导;不同血清型rAAV对NK92细胞的转导效率由高到低依次为rAAV6、rAAV2和rAAV9;用金雀异黄酮预处理NK92细胞,能够显著提高病毒转导效率,而其他促进剂的添加对病毒转导效率无显著影响。通过上述条件的优化,显著提高了rAAV对NK92细胞的转导效率,为rAAV载体介导工程化NK92细胞奠定基础。

To improve the transduction efficiency of recombinant adeno-associated virus(rAAV)in NK92 cells,the number of cells,concentration of IL-2 in the medium,and serotype and dosage of rAAV were explored to optimize cell state and viral transfection conditions.Then,zinc chloride(ZnCl_(2)),chloroquine,polyvinyl alcohol(PVA)and genistein with different concentration were added separately during transfection to further improve the viral transduction efficiency.The results showed that,at cell number of 5×10^(5),the expression efficiency of enhanced green fluorescent protein(EGFP)was relatively high.When the IL-2 concentration was 1000 IU/mL,NK92 cells were most suitable for virus transfection.The transduction efficiency of different serotypes of rAAV in NK92 cells was rAAV6,rAAV2 and rAAV9 in descending order.Pretreatment of NK92 cells with genistein could significantly increase the viral transduction efficiency,while the addition of other reagents had no significant effect.Through the optimization of the above conditions,the transduction efficiency of rAAV to NK92 cells could be significantly improved,which provided evidence for functional genetic modification of NK92 cells by rAAV.