Molecular basis of giant Extracellular Matrix Binding Protein (Embp) mediated Staphylococcus epidermidis adherence to fibronectin and biofilm accumulation on artificial surfaces

该项目的总体目标是全面了解Embp介导的表皮葡萄球菌生物膜形成的分子基础及其在实验异物相关感染发病过程中与细菌/宿主相互作用的相关性。通过使用各种实验方法，包括分子生物学技术、细胞培养系统和建立种植体相关感染的三维模型系统，解决Embp依赖性表皮葡萄球菌毒力的关键方面，将达到目标。1. 我们将在分子/生化水平上描述Embp / FN相互作用的确切模式，并分析它们在表皮葡萄球菌与表面组织FN结合过程中的作用。此外，还将分析FN纤原形成过程中动态构象变化的重要性以及连续暴露的隐表位，如FN III12至14。理由：表皮葡萄球菌与FN结合对植入物相关感染的发生至关重要。事实证明，Embp对于依赖FN的表皮葡萄球菌粘附在种植体表面是必要的。然而，这种相互作用的确切分子决定因素目前尚不清楚。我们小组的工作表明，Embp采用迄今未知的与FN相互作用的模式，包括结合到隐模FN III12至14。2. 将建立并验证种植体相关感染的三维模型。该模型将允许分析Embp依赖性表皮葡萄球菌植入物定植、生物膜形成以及与宿主先天免疫细胞相互作用期间的时空动态。合理：目前用于分析表皮葡萄球菌生物膜形成的分析系统不能反映出发生在外来物质和周围组织(由细胞外基质和多种常驻细胞（如成纤维细胞、巨噬细胞）组成的界面上的体内感染的复杂情况。这种特殊的地形，基本上适用于大多数表皮葡萄球菌装置相关感染（例如假体关节、脑脊液分流、人工心脏瓣膜），可能对表皮葡萄球菌的生物膜发育和与宿主细胞的动态相互作用产生巨大影响。总的来说，对这些目标的分析不仅将为细菌发病机制和微生物/宿主相互作用的一般方面提供新的见解，而且更具体地解决与表皮葡萄球菌植入物感染发病机制相关的核心方面。考虑到临床问题的严重性，拟议的项目可以为开发新的治疗和/或预防方法提供新的起点，这些方法是对抗破坏性设备相关的表皮葡萄球菌感染所必需的。

The overall objective of this project is to obtain comprehensive insights into the molecular basis of Embp mediated S. epidermidis biofilm formation and its relevance to bacterial / host interactions during pathogenesis of experimental foreign material associated infections. The objective will be reached by addressing key aspects of Embp dependent S. epidermidis virulence using a variety of experimental approaches, including molecular biological techniques, cell culture systems and establishment of three dimensional model systems of implant associated infections. 1. We will characterize the exact modalities of Embp / FN interactions on a molecular / biochemical level and analyze their role during S. epidermidis binding to surface organized FN. In addition, the importance of dynamic conformational changes during FN fibrillogenesis and consecutive exposure of cryptic epitopes, e.g. FN III12 to 14, will be analyzed.Rational: S. epidermidis binding to FN is of paramount importance for initiation of implant associated infections. Embp proved to be necessary for FN dependent S. epidermidis adherence to implant surfaces. However, the exact molecular determinants of this interaction are currently unclear. Work from our groups demonstrates that Embp employs a so far unknown mode of interaction with FN involving binding to cryptic modules FN III12 to 14. 2. A three dimensional model of implant associated infections will be established and validated. The model will allow for analysis of the temporal and spatial dynamics during Embp dependent S. epidermidis implant colonization, biofilm formation and interactions with host innate immune cells. Rational: Currently assay systems used for analysis of S. epidermidis biofilm formation do not reflect the complex setting of an in vivo infection occurring at the interface between foreign materials and the surrounding tissues consisting of extracellular matrix, and a diverse set of resident cells (e.g. fibroblasts, macrophages). This specific topography, which essentially applies for most S. epidermidis device related infections (e.g. associated with prosthetic joints, CSF shunts, artificial heart valves) could have dramatic impact on biofilm development and dynamic interactions of S. epidermidis with host cells.Collectively, analysis of these objectives will not only provide novel insights into general aspects of bacterial pathogenesis and microbe / host interactions, but more specifically address central aspects related to the pathogenesis of S. epidermidis implant infections. Considering the gravity of the clinical problem, the proposed project could provide new starting points for development of novel therapeutic and / or prophylactic approaches that are necessary to combat devastating device related S. epidermidis infections.