Human Neutrophil Response to Trichomonas vaginalis

项目摘要/摘要 阴道毛滴虫(TV)是一种原生动物病原体，可导致最常见的非病毒性 在美国和世界范围内传播感染。TV的致病机制和免疫机制的研究进展 有限的。然而，电视抗药性菌株的上升，与新兴的欣赏之间的联系 无症状的电视感染与炎症驱动的病理需要更好地理解这一点 流行的人类感染。值得注意的是，电视感染与生殖并发症有关，增加了 对艾滋病毒的易感性、宫颈癌发病率的增加以及前列腺癌侵袭性的增加。 临床观察、前人的工作和我们实验室的初步研究都指向中性粒细胞 (PMN)作为反电视免疫的关键人物。PMN是极具破坏性的细胞，它会传播一系列 组织损伤和炎症，这一直与炎症病理有关，启动新的 癌症和现有肿瘤的恶化。如果感染没有得到有效的清除， 炎症可能会长期存在。虽然PMN在电视感染中的重要性已经被认识到，但他们如何 抗击电视感染的工作并不是其特点。因此，我们的目标是(I)确定PMN如何扼杀电视，以及(Ii) 确定在TV感染过程中PMN在炎症形成中的作用。我们的初步数据表明 PMN使用巨噬细胞增多症(trogo=nibble)杀死电视，这是一种效应细胞从 目标单元格。我们将从人血中分离PMN，并与电视共培养。然后我们将使用流式细胞术- 基于细胞溶解试验、活细胞成像显微镜和成像流式细胞术来确定PMN 巨噬细胞增多症导致电视节目死亡。我们还将确定PMN是否使用中性粒细胞胞外陷阱 杀死电视的蚊虫，PMN向外释放核内容物以诱捕、固定和杀死的过程 病原体。为了评估蚊虫感染，我们将使用细胞外DNA定量、成像和流式细胞术。 以细胞溶解试验为基础。此外，为了确定PMN-TV相互作用对炎症的影响， 我们将探索PMN-TV共培养上清液，以确定电视诱导的细胞因子程序。我们会 使用直播电视与热灭活电视来确定电视介导的组织破坏在 炎症，我们还将在阴道上皮细胞存在的情况下进行TV-PMN共培养 确定电视和中性粒细胞对上皮层的破坏如何导致炎症。《知识》 从这项关于中性粒细胞如何清除急性TV感染以及炎症信号是什么的研究中获得 建立，将有助于提出潜在的战略，以监测炎症病理，宫颈癌和 与电视感染相关的前列腺癌，并为潜在免疫治疗干预措施的设计提供信息。

PROJECT SUMMARY / ABSTRACT Trichomonas vaginalis (Tv) is a protozoan pathogen that causes the most common non-viral sexually transmitted infection in the United States and worldwide. Studies on Tv pathogenesis and immunity have been limited. However, the rise in drug resistant strains of Tv, and an emerging appreciation of the link between asymptomatic Tv infections with inflammation-driven pathologies demands better understanding of this prevalent human infection. Notably, Tv infection has been linked to reproductive complications, increased susceptibility to HIV, increased incidence of cervical cancer, and increased aggressiveness of prostate cancer. Clinical observation, previous work by others, and preliminary studies in our laboratory point to neutrophils (PMN) as the key player in anti-Tv immunity. PMN are extremely destructive cells, propagating a cascade of tissue damage and inflammation, which has been associated with inflammatory pathologies, initiation of new cancers, and exacerbation of existing neoplasms. If an infection is not cleared efficiently, the cycle of inflammation could persist chronically. While the importance of PMN in Tv infection is recognized, how they work to fight Tv infection is not characterized. Therefore, we aim to (i) determine how PMN kill Tv, and (ii) determine the role of PMN in establishing inflammation during Tv infection. Our preliminary data suggest that PMN kill Tv using trogocytosis (trogo= nibble), a process by which effector cells take small “bites” from target cells. We will isolate PMN from human blood and co-culture with Tv. We will then use flow cytometry- based cytolysis assays, live cell imaging microscopy and imaging flow cytometry to determine whether PMN trogocytosis leads to death of Tv. We will also determine whether PMN use neutrophil extracellular traps NETosis to kill Tv, a process by which PMN release nuclear contents externally to ensnare, immobilize and kill pathogens. To assess NETosis, we will use extracellular DNA quantification, imaging, and flow-cytometry based cytolysis assays. In addition, to determine the consequences of PMN-Tv interaction on inflammation, we will probe supernatants of PMN-Tv co-cultures to determine the cytokine program that Tv elicits. We will use live Tv versus heat-inactivated Tv to determine the contribution of Tv-mediated tissue destruction on inflammation, and we will also perform Tv-PMN co-cultures in the presence of vaginal epithelial cells to determine how destruction of the epithelial layer by Tv and PMN contributes to inflammation. The knowledge gained from this study on how acute Tv infection is cleared by PMN and what inflammatory signals are established, will help to suggest potential strategies to monitor inflammatory pathologies, cervical cancers and prostate cancers associated with Tv infection, and inform the design of potential immunotherapy interventions.