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Abstract：
Colonization of the endometrium by pathogenic bacteria ascending from the lower female reproductive tract (FRT) is associated with many gynecologic and obstetric health complications. To study these host-microbe interactions in vitro, we developed a human three-dimensional (3-D) endometrial epithelial cell (EEC) model using the HEC-1A cell line and the rotating wall vessel (RWV) bioreactor technology. Our model, composed of 3-D EEC aggregates, recapitulates several functional/structural characteristics of human endometrial epithelial tissue, including cell differentiation, the presence of junctional complexes/desmosomes and microvilli, and the production of membrane-associated mucins and Toll-like receptors (TLRs). TLR function was evaluated by exposing the EEC aggregates to viral and bacterial products. Treatment with poly(I·C) and flagellin but not with synthetic lipoprotein (fibroblast-stimulating lipoprotein 1 [FSL-1]) or lipopolysaccharide (LPS) significantly induced proinflammatory mediators in a dose-dependent manner. To simulate ascending infection, we infected EEC aggregates with commensal and pathogenic bacteria: Lactobacillus crispatus, Gardnerella vaginalis, and Neisseria gonorrhoeae All vaginal microbiota and N. gonorrhoeae efficiently colonized the 3-D surface, localizing to crevices of the EEC model and interacting with multiple adjacent cells simultaneously. However, only infection with pathogenic N. gonorrhoeae and not infection with the other bacteria tested significantly induced proinflammatory mediators and significant ultrastructural changes to the host cells. The latter observation is consistent with clinical findings and illustrated the functional specificity of our system. Additionally, we highlighted the utility of the 3-D EEC model for the study of the pathogenesis of N. gonorrhoeae using a well-characterized ΔpilT mutant. Overall, this study demonstrates that the human 3-D EEC model is a robust tool for studying host-microbe interactions and bacterial pathogenesis in the upper FRT.
摘要：
从女性下生殖道（FRT）上升的病原菌对子宫内膜的定植与许多妇科和产科健康并发症有关。为了在体外研究这些宿主-微生物相互作用，我们使用HEC-1A细胞系和旋转壁血管（RWV）生物反应器技术开发了人类三维（3-D）子宫内膜上皮细胞（EEC）模型。我们的模型，由3-DEEC骨料组成，概括了人类子宫内膜上皮组织的几种功能/结构特征，包括细胞分化，连接复合物/桥粒和微绒毛的存在，以及膜相关粘蛋白和Toll样受体（TLR）的产生。通过将EEC聚集体暴露于病毒和细菌产物来评估TLR功能。用聚（I·C）和鞭毛蛋白治疗，但不使用合成脂蛋白（成纤维细胞刺激脂蛋白1[FSL-1]）或脂多糖（LPS）治疗，以剂量依赖性方式显着诱导促炎介质。为了模拟上升感染，我们用共生和致病菌感染了EEC聚集体：crispatus乳杆菌，阴道加德纳菌,和淋病奈瑟菌所有阴道微生物群和淋病奈瑟菌有效地定植在3-D表面，定位到EEC模型的缝隙，并同时与多个相邻细胞相互作用。然而,仅感染致病性淋病奈瑟菌，而不感染其他测试细菌，显着诱导促炎介质和宿主细胞的显着超微结构变化。后一种观察结果与临床发现一致，并说明了我们系统的功能特异性。此外,我们强调了3-DEEC模型在使用特征明确的ΔpilT突变体研究淋病奈瑟菌发病机理中的实用性。总的来说,这项研究表明，人类3-DEEC模型是研究上FRT中宿主-微生物相互作用和细菌发病机理的强大工具。
