Department of Molecular and Clinical Medicine/Wallenberg Laboratory, University of Gothenburg, Göteborg, Sweden.
Gene Ther. 2012 Nov;19(11):1041-7. doi: 10.1038/gt.2011.195. Epub 2011 Dec 1.
In gene therapy, tissue-specific promoters are useful tools to direct transgene expression and improve efficiency and safety. Macrophage-specific promoters (MSPs) have previously been published using different delivery systems. In this study, we evaluated five different MSPs fused with green fluorescent protein (GFP) to delineate the one with highest specificity using lentiviral delivery. We compared three variants of the CD68 promoter (full length, the 343-bp proximal part and the 150-bp proximal part) and two variants (in forward and reverse orientation) of a previously characterized synthetic promoter derived from elements of transcription factor genes. We transduced a number of cell lines and primary cells in vitro. In addition, hematopoietic stem cells were transduced with MSPs and transferred into lethally irradiated recipient mice. Fluorescence activated cell sorting analysis was performed to determine the GFP expression in different cell populations both in vitro and in vivo. We showed that MSPs can efficiently be used for lentiviral gene delivery and that the 150-bp proximal part of the CD68 promoter provides primarily macrophage-specific expression of GFP. We propose that this is the best currently available MSP to use for directing transgene expression to macrophage populations in vivo using lentiviral vectors.
在基因治疗中，组织特异性启动子是将转基因表达导向特定组织并提高效率和安全性的有用工具。先前已经使用不同的递送系统发布了巨噬细胞特异性启动子（MSP）。在这项研究中，我们评估了与绿色荧光蛋白（GFP）融合的五种不同的 MSP，以使用慢病毒递送系统来确定特异性最高的一种。我们比较了 CD68 启动子的三个变体（全长、343bp 近端部分和 150bp 近端部分）和两个变体（正向和反向取向），这两个变体是先前从转录因子基因元件中鉴定出的合成启动子。我们在体外转导了许多细胞系和原代细胞。此外，造血干细胞用 MSP 转导并转移到致死性辐射的受体小鼠中。通过荧光激活细胞分选分析，确定了不同细胞群体在体外和体内的 GFP 表达。我们表明 MSP 可有效地用于慢病毒基因递送，并且 CD68 启动子的 150bp 近端部分主要提供 GFP 的巨噬细胞特异性表达。我们建议，这是目前使用慢病毒载体在体内将转基因表达导向巨噬细胞群体的最佳 MSP。
