Papageorgiou Konstantina, Isenberg David A, Latchman David S
Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK.
J Immunol Methods. 2002 Dec 15;270(2):235-46. doi: 10.1016/s0022-1759(02)00299-5.
Peripheral blood mononuclear cells (PBMCs) represent a significant target for gene delivery both for therapeutic and experimental purposes. Thus far however, it has proved difficult to develop vectors capable of high efficient gene delivery to unstimulated PBMCs. We have tested a range of different vectors derived from herpes simplex virus (HSV) which differ in their degree of disablement in terms of their gene delivery efficiency to unstimulated human PBMCs and ability to deliver a reporter gene. None of the viruses had any significant toxic effect in PBMCs. However, optimal gene delivery to unstimulated PBMCs was obtained with a semidisabled virus lacking functional genes encoding ICP34.5 and Vmw65 which was more efficient than either nondisabled or more extremely disabled viruses. Expression of green fluorescent protein (GFP) with this virus was observed in up to 50% of PBMCs 1 day after infection, and reporter gene expression was detectable by Western blotting and immunofluorescence at undiminished levels at the longest time points tested, up to 5 days after infection. This optimised HSV vector may thus represent an effective tool for gene delivery to unstimulated PBMCs in culture.
外周血单个核细胞（PBMCs）是治疗和实验目的基因递送的重要靶点。然而，到目前为止，开发能够高效地将基因递送至未激活的PBMCs的载体已被证明是困难的。我们测试了一系列源自单纯疱疹病毒（HSV）的不同载体，这些载体在向未激活的人PBMCs递送基因的效率以及递送报告基因的能力方面，其失活程度有所不同。这些病毒在PBMCs中均无明显毒性作用。然而，对于未激活的PBMCs，使用一种缺乏编码ICP34.5和Vmw65功能基因的半失活病毒可实现最佳的基因递送，该病毒比未失活或极度失活的病毒更有效。感染后1天，使用这种病毒在高达50%的PBMCs中观察到绿色荧光蛋白（GFP）的表达，并且在最长测试时间点（感染后5天）通过蛋白质印迹法和免疫荧光法可检测到报告基因的表达，且表达水平未降低。因此，这种优化的HSV载体可能是向培养中的未激活PBMCs递送基因的有效工具。
