In vivo Formation and Maintenance of Salmonella-Containing Vacuoles (SCVs) in Enterocytes using a Novel Neonatal Mouse Model

人类病原体肠道沙门氏菌对非吞噬细胞的侵袭以及操纵内体隔室以允许细胞内细菌存活和增殖已在体外进行了广泛的研究。然而，由于缺乏合适的动物模型，尚未对非吞噬细胞（例如上皮细胞）中沙门氏菌液泡（SCV）的建立进行体内研究。基于建立一种新的新生小鼠感染模型，可以分析体内肠上皮细胞的侵袭和 SCV 的形成，本研究项目旨在表征感染过程中原代肠上皮细胞的内体区室。我们将定义防止膜破裂的宿主和细菌因素，并有助于 SCV 形成和上皮内运输，以促进细菌增殖和粘膜易位。计划的研究项目解决以下问题：（i）体内完全分化的肠上皮细胞和感染的类器官共培养物中含有沙门氏菌的空泡（SCV）的详细表征。 (ii) 分析沙门氏菌效应蛋白对 SCV 生成和极化原代肠细胞内囊泡运输的作用。(iii) 鉴定有助于 SCV 形成或介导内体破坏和细菌杀伤的宿主衍生信号。 我们期望对体内肠上皮细胞中含有沙门氏菌的空泡的建立的首次分析将揭示细胞内宿主-微生物相互作用和沙门氏菌感染发病机制的新的重要见解。

Invasion of non-phygocytic cells by the human pathogen Salmonella enterica and the manipulation of the endosomal compartment to allow intracellular bacterial survival and proliferation have been studied extensively in vitro. However, establishment of the Salmonella-containing vacuole (SCV) in non-phagocytic cells such as epithelial cells has not been investigated in vivo due to the lack of a suitable animal model. Based on the establishment of a new infection model of neonate mice that allows the analysis of enterocyte invasion and SCV formation in vivo, the present research project aims at characterizing this endosomal compartment in primary enterocytes during the course of infection. We will define host and bacterial factors that prevent membrane disruption, and contribute to SCV formation and intraepithelial trafficking to facilitate bacterial proliferation and mucosal translocation.The planned research project addresses the following issues:(i) Detailed characterization of Salmonella-containing vacuoles (SCV) in fully differentiated enterocytes in vivo and in infected organoid co-cultures. (ii) Analysis of the role of Salmonella effector proteins for SCV generation and vesicular trafficking within polarized primary enterocytes.(iii) Identification of host-derived signals that contribute to SCV formation or mediate endosomal disruption and bacterial killing. We expect that this first analysis of the establishment of Salmonella-containing vacuoles in enterocytes in vivo will reveal new and important insight in the intracellular host-microbial interaction and pathogenesis of Salmonella infection.