Development of a novel, highly quantitative in vivo model for the study of biofilm-impaired cutaneous wound healing
Development of a novel, highly quantitative in vivo model for the study of biofilm-impaired cutaneous wound healing
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DOI：
10.1111/j.1524-475x.2011.00690.x
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发表时间：
2011-05-01
影响因子：
2.9
通讯作者：
Mustoe, Thomas A.
中科院分区：
文献类型：
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作者：
Gurjala, Anandev N.;Geringer, Matthew R.;Mustoe, Thomas A.
越来越多的证据表明，除了缺氧，缺血-再灌注损伤，和内在的宿主因素，细菌生物膜代表慢性伤口发病机制的第四个主要支柱。鉴于迄今为止大多数研究依赖于体外或观察性临床数据，我们的目的是开发一种新的定量动物模型，从而能够进一步研究体内生物膜假说。在新西兰兔耳中产生皮肤穿孔伤口，并用作未感染的对照，或用绿色荧光蛋白标记的金黄色葡萄球菌接种以形成具有主要处于增殖期或生物膜期的细菌的伤口。扫描电子显微镜和荧光显微镜观察表明，S。金黄色葡萄球菌在接种24小时内在伤口中快速形成成熟的生物膜，通过在创伤和接种后的不同时间点的连续细菌计数测量和激光扫描共聚焦成像，可以看到生物膜活力随时间的持续。炎症标志物证实，生物膜表型产生特征性的、持续的、低度炎症反应，并且随着时间的推移，生物膜损害上皮迁移和肉芽组织向内生长，如组织学所示。我们已经建立并验证了一个高度定量的，可重复的体内生物膜模型，同时提供证据表明，生物膜表型特别有助于深刻的皮肤伤口愈合障碍。我们的模型强调了细菌生物膜在慢性伤口发病机制中的重要性，为进一步探究细菌生物膜-宿主相互作用的基础生物学和生物膜治疗的高通量测试提供了体内平台。
A growing body of evidence suggests that in addition to hypoxia, ischemia-reperfusion injury, and intrinsic host factors, bacterial biofilms represent a fourth major pillar in chronic wound pathogenesis. Given that most studies to date rely on in vitro or observational clinical data, our aim was to develop a novel, quantitative animal model enabling further investigation of the biofilm hypothesis in vivo. Dermal punch wounds were created in New Zealand rabbit ears, and used as uninfected controls, or inoculated with green fluorescent protein-labeled Staphylococcus aureus to form wounds with bacteria predominantly in the planktonic or biofilm phase. Epifluorescence and scanning electron microscopy revealed that S. aureus rapidly forms mature biofilm in wounds within 24 hours of inoculation, with persistence of biofilm viability over time seen through serial bacterial count measurement and laser scanning confocal imaging at different time points postwounding and inoculation. Inflammatory markers confirmed that the biofilm phenotype creates a characteristic, sustained, low-grade inflammatory response, and that over time biofilm impairs epithelial migration and granulation tissue in-growth, as shown histologically. We have established and validated a highly quantitative, reproducible in vivo biofilm model, while providing evidence that the biofilm phenotype specifically contributes to profound cutaneous wound healing impairment. Our model highlights the importance of bacterial biofilms in chronic wound pathogenesis, providing an in vivo platform for further inquiry into the basic biology of bacterial biofilm-host interaction and high-throughput testing of antibiofilm therapeutics.