Direct injection of plasmid DNA into the skin induces dermatitis by activation of monocytes through toll-like receptor 9
Direct injection of plasmid DNA into the skin induces dermatitis by activation of monocytes through toll-like receptor 9
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DOI：
10.1002/jgm.709
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发表时间：
2005-05-01
影响因子：
3.5
通讯作者：
Shimizu, H
中科院分区：
文献类型：
--
作者：
Sawamura, D;Abe, R;Shimizu, H
在体内，将DNA直接注射到皮肤中可以有效地将基因转移到角质形成细胞中。然而，已知细菌DNA是脊椎动物免疫细胞和免疫系统的有效刺激。针对该方法的临床应用，本研究考察了直接注射质粒DNA是否会引起皮肤不良反应。方法制备多种质粒制剂，直接注射于大鼠和人皮肤。通过迁移、IL-6生成和活性氧生成实验来确定参与反应的原代细胞类型。以tlr9敲除小鼠为实验对象，研究toll样受体（TLR） 9的参与情况。结果将几种质粒制剂注射到大鼠和人皮肤后，在处理部位产生炎症反应。质粒制剂中内毒素的污染使皮肤炎症反应恶化。免疫组化分析显示，皮损内浸润细胞以单核细胞和中性粒细胞为主。进一步研究迁移、IL-6产生和活性氧产生的实验表明，主要负责的细胞是单核细胞而不是中性粒细胞。由于最近发现胞嘧啶-鸟苷二核苷酸（CpG）基序在细菌DNA的免疫反应诱导中起关键作用，并且细胞反应是由TLR9介导的，因此我们将质粒注射到TLR9敲除小鼠的耳皮肤中。与对照组相比，敲除小鼠的耳部肿胀减少，表明质粒dna诱导的皮炎主要由TLR9介导。结论注射质粒DNA可诱导单核细胞通过TRL9激活引起的皮肤炎症。因此，在临床使用裸DNA注射方法时，我们应该尝试对抗这种皮炎。版权所有(c) 2005 John Wiley & Sons, Ltd。
Background Direct injection of naked DNA into skin can be efficiently used to transfer genes into keratinocytes in vivo. However, bacterial DNA is known to be a potent stimulus for vertebrate immune cells and immune systems. Towards the clinical use of this method, this study examined whether the application of plasmid DNA by direct injection induces any adverse skin effects.Methods Several plasmid preparations were prepared and directly injected into rat and human skin. Migration, IL-6 production, and reactive oxygen production assays were performed to determine the type of the primary cells responsible for the reaction. Involvement of toll-like receptor (TLR) 9 was examined by experiments using TLR9-knockout mice.Results Injection of several plasmid preparations into rat and human skin resulted in an inflammatory reaction at the treated site. Contamination by endotoxin in the plasmid preparation was shown to worsen this skin inflammation reaction. Immunohistochemical analysis showed that the infiltrating cells in the skin lesions were predominantly monocytes and neutrophils. Further experiments examining migration, IL-6 production, and reactive oxygen production indicated that the primary responsible cells were monocytes rather than neutrophils. Since it was recently shown that cytosine-guanosine dinucleotide (CpG) motif is critical for immune reaction induction in bacterial DNA and cellular responses were mediated by TLR9, we injected plasmids into the ear skin of TLR9-knockout mice. A decrease in ear swelling was noted in the knockout mice, compared to controls, suggesting that plasmid-DNA-induced dermatitis was mediated mostly by TLR9.Conclusions This study demonstrates that injection of plasmid DNA induces skin inflammation initiated by monocyte activation via TRL9. We should therefore attempt to counteract this dermatitis during the clinical use of the naked DNA injection method in skin. Copyright (c) 2005 John Wiley & Sons, Ltd.