Antimicrobial peptides and pro-inflammatory cytokines are differentially regulated across epidermal layers following bacterial stimuli
Antimicrobial peptides and pro-inflammatory cytokines are differentially regulated across epidermal layers following bacterial stimuli
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DOI：
10.1111/exd.12259
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发表时间：
2013-12-01
影响因子：
3.6
通讯作者：
Follet-Gueye, Marie-Laure
中科院分区：
文献类型：
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作者：
Percoco, Giuseppe;Merle, Chloe;Follet-Gueye, Marie-Laure
皮肤是身体和环境之间的天然屏障，并被大量微生物定殖。在这里，我们报告一个完整的分析人类皮肤外植体微生物刺激的反应。使用这种离体模型，我们在基因和蛋白质水平上分析了表皮细胞对表皮葡萄球菌（S.epidermidis）和荧光假单胞菌（P.fluorescens）的反应，这些细菌存在于皮肤微生物群中。我们发现，这两种细菌影响皮肤外植体的结构，而不穿透活表皮。我们通过实时定量聚合酶链反应（qPCR）显示，表皮葡萄球菌和荧光假单胞菌增加了编码抗菌肽（AMP）的转录物水平，包括人防御素（hBD）2和hBD 3，以及促炎细胞因子白细胞介素（IL）-1和（IL）-1-，以及IL-6。此外，我们使用激光捕获显微切割（LCM）结合qPCR分析了细菌刺激对与先天免疫和表皮层炎症反应相关的基因表达谱的影响。我们发现AMP转录本主要在基底上角质形成细胞中上调。相反，促炎细胞因子的表达在下表皮中上调。这些发现证实了蛋白定位使用特异性抗体耦合到光学或电子显微镜。这项工作强调了使用LCM对人类皮肤外植体模型进行进一步研究的潜在价值，以研究表皮微生物群对人类皮肤生理学的作用和影响。
The skin is a natural barrier between the body and the environment and is colonised by a large number of microorganisms. Here, we report a complete analysis of the response of human skin explants to microbial stimuli. Using this ex vivo model, we analysed at both the gene and protein level the response of epidermal cells to Staphylococcus epidermidis (S.epidermidis) and Pseudomonas fluorescens (P.fluorescens), which are present in the cutaneous microbiota. We showed that both bacterial species affect the structure of skin explants without penetrating the living epidermis. We showed by real-time quantitative polymerase chain reaction (qPCR) that S.epidermidis and P.fluorescens increased the levels of transcripts that encode antimicrobial peptides (AMPs), including human defensin (hBD)2 and hBD3, and the pro-inflammatory cytokines interleukin (IL)-1 and (IL)-1-, as well as IL-6. In addition, we analysed the effects of bacterial stimuli on the expression profiles of genes related to innate immunity and the inflammatory response across the epidermal layers, using laser capture microdissection (LCM) coupled to qPCR. We showed that AMP transcripts were principally upregulated in suprabasal keratinocytes. Conversely, the expression of pro-inflammatory cytokines was upregulated in the lower epidermis. These findings were confirmed by protein localisation using specific antibodies coupled to optical or electron microscopy. This work underscores the potential value of further studies that use LCM on human skin explants model to study the roles and effects of the epidermal microbiota on human skin physiology.