Microbially competent 3D skin - a novel test system for studying host-microbe interaction

The skin`s microbiome is predominantly commensalic, harbouring a metabolic potential far exceeding that of its host. While toxicologically relevant there is still a lack of suitable models.  We now report on a new biologically characterised co-culture that allows studying microbe-host interactions for extended periods of time in situ . The system is based on a commercially available 3D skin model. In a proof of concept this model was colonised with single and mixed cultures of two selected skin commensals. Two different methods were used to quantify the bacteria on the surface of the skin models. While M. luteus established a stable co-culture, P. oleovorans maintained slow continuous growth over the 8 day cultivation period. A detailed skin transcriptome analysis showed bacterial colonisation leading to up to 3318 significant changes. Additionally FACS, ELISA and Western blot analyses were carried out to analyse secretion of cytokines and growth factors. Changes found in colonised skin were varied dependent on the bacterial species used and comprised immunomodulatory functions, such as secretion of IL-1α/β, Il-6, antimicrobial peptides and increased gene transcription of IL-10 and TLR2. The colonisation also influenced the secretion of many grows factors as VFGFA and FGF2. Notably, many of these changes have already previously been associated with the presence of skin commensals. Concomitantly the model gained first insights on the microbiome’s influence on skin xenobiotic metabolism (i.e., CYP1A1, CYP1B1 and CYP2D6) and olfactory receptor expression. The system provides urgently needed experimental access for assessing the toxicological impact of the microbiome’s xenobiotic metabolism in situ. Development of a microbial competent skin model and its characterization. For this purpose we analyzed transcription of co-cultures of EpiDermFT™ (MatTek) skin tissue and two different skin isolates (M. luteus and P. oleovorans) using the Affymetrix Human Clariom S. Array data was processed by the company ATLAS Biolabs GmbH (Friedrichstraße 147, 10117 Berlin, Germany). Three biological replicates were performed for each condition.

皮肤微生物组以共生菌为主，其代谢潜力远超宿主。尽管在毒理学领域具有重要研究价值，但目前仍缺乏合适的研究模型。本研究报道了一种经过生物学表征的新型共培养体系，可用于原位（in situ）研究微生物与宿主的长期相互作用。该体系基于商业化的三维皮肤模型。在概念验证实验中，研究人员使用两种筛选出的皮肤共生菌的单培养及混合培养物对该模型进行定植。研究采用两种不同方法对皮肤模型表面的细菌进行定量。藤黄微球菌（M. luteus）可形成稳定的共培养体系，而食油假单胞菌（P. oleovorans）在8天的培养周期中维持缓慢的持续生长。详细的皮肤转录组分析显示，细菌定植可导致最多3318个基因表达发生显著变化。此外，研究还通过流式细胞术（FACS）、酶联免疫吸附实验（ELISA）及蛋白质印迹（Western blot）分析了细胞因子与生长因子的分泌情况。定植后皮肤的变化因所使用的细菌种类而异，涵盖免疫调节功能相关变化，例如IL-1α/β、IL-6的分泌、抗菌肽的产生，以及IL-10和TLR2的基因转录水平上调。定植还影响了包括血管内皮生长因子A（VEGFA）和成纤维细胞生长因子2（FGF2）在内的多种生长因子的分泌。值得注意的是，其中许多变化此前已被证实与皮肤共生菌的存在相关。与此同时，该模型首次揭示了微生物组对皮肤异生物质代谢（即CYP1A1、CYP1B1及CYP2D6）及嗅觉受体表达的影响。该体系为原位评估微生物组的异生物质代谢所产生的毒理学影响提供了亟需的实验手段。本研究开发了一种具备微生物定植能力的皮肤模型并对其进行表征。为此，研究人员使用Affymetrix Human Clariom S芯片对EpiDermFT™ (MatTek)皮肤组织与两种皮肤分离株（M. luteus和P. oleovorans）的共培养物进行转录组分析。芯片数据由德国柏林ATLAS Biolabs GmbH公司（Friedrichstraße 147, 10117 Berlin, Germany）进行处理。每个实验条件均设置3次生物学重复。