EHRLICHIA-GRANULOCYTE INTERACTIONS AND INFECTION

描述(改编自申请者摘要)：人粒细胞 埃立克次体病(Hge)是一种新近出现的由硬蜱传播的传染病。 在过去的十年里，这种情况越来越频繁地被报道。这个 病原体被归类在埃利希氏基因组II和 被认为是吞噬埃利希氏菌的变种或亚种。虽然 许多--也许大多数--人类感染要么是亚临床感染，要么是可解决的 在没有诊断的情况下，严重疾病确实会导致长时间的住院治疗 重症监护，并可进展至死亡。人类的表现形式 这种疾病没有特异性，但可能表现为一种中毒性休克样疾病，成人 呼吸窘迫综合征(ARDS)和死亡病例与 机会性感染。据报道，类似的关联也与 感染吞噬埃希氏菌群生物的家畜。自然的 靶细胞主要是中性粒细胞，尽管早幼粒细胞可以是 在体外感染和髓系祖细胞被假设为一个部位 体内感染的可能性。相反，巨噬细胞和单核细胞样细胞的感染 细胞系会流产，生物体就会被杀死。有趣的是， 人类疾病的严重程度并不反映高循环埃利希菌 装填。先前来自感染吞噬埃希氏菌的动物的数据表明 中性粒细胞功能受损--包括趋化、吞噬和杀伤。 这些“失活”表型可能与 与内皮细胞发生结合并最终导致 内皮细胞释放促炎细胞因子，尤其是 巨噬细胞，导致中毒性休克样损伤的触发。 PI提出了以下假设：1)吞噬细胞性埃立克体群 与CD15相关的粒细胞表面受体结合并启动内吞作用 通过它们的主要表面蛋白(MSP)抗原；2)结合、内化， 和吞噬E.phila群Ehrlichiae启动繁殖 细胞因子/趋化因子在粒细胞中的表达及变化 激活/失活与功能；以及3)E.吞噬细胞组 埃立克次体通过表达多种蛋白潜在地影响宿主细胞的功能 直接与宿主细胞中的DNA调控成分相互作用 染色体。这些假设将通过五个具体目标进行测试：i) 展示埃立克体与宿主细胞膜相互作用的形态 HL60细胞处于不同分化状态；ii)鉴定E。 吞噬细胞型MSPs作为黏附素及CD15相关分子的作用 作为埃立克体附着和进入的主要表面受体的结构 细胞；三)表征细菌结合和内化的影响 对宿主粒细胞表面黏附分子的表达， 粒细胞-内皮细胞黏附，吞噬活性， 激活/失活、微生物杀伤和细胞因子/趋化因子的表达； 以及iv)确定吞噬E.phila组ankyrin的染色体配基 蛋白质，EPANK1。

DESCRIPTION (Adapted from the Applicant's Abstract): Human Granulocytic Ehrlichiosis (HGE) is a recently emergent tick-transmitted infectious disease that has been reported with increasing frequency during the last decade. The causative agent is classified within the ehrlichial genogroup II and is considered to be a variant or subspecies of Ehrlichia phagocytophila. Although many and perhaps most of the human infections are either subclinical or resolve without diagnosis, severe disease does result in prolonged hospitalization with intensive care and can progress to cause death. The manifestations of human disease are not specific but may manifest as a toxic shock-like illness, adult respiratory distress syndrome (ARDS), and fatal cases have an association with opportunistic infections. A similar association has been reported with the domestic animals infected by E. phagocytophila group organisms. The natural target cell is primarily neutrophils, although premyelocytic cells can be infected in vitro and myeloid progenitors have been hypothesized to be a site of infection in vivo. In contrast, infection of macrophages and monocyte-like cell lines is abortive and the organisms are killed. Interestingly, the severity of disease in humans does not reflect a high circulating ehrlichial load. Previous data from animals infected with E. phagocytophila indicates impaired neutrophil function - including chemotaxis, phagocytosis, and killing. These "deactivation" phenotypes may be concomitantly associated with "activation" in which binding to endothelial cells occurs and culminates in release of proinflammatory cytokines by endothelial cells and especially macrophages, leading to the triggering of a toxic-shock like injury. The PI proposes the following hypotheses: 1) E. phagocytophila group ehrlichiae bind to CD15-associated granulocyte surface receptors and initiate endocytosis via their major surface protein (MSP) antigens; 2) Binding, internalization, and propagation of E. phagocytophila group ehrlichiae initiate cytokine/chemokine expression and changes in granulocyte activation/inactivation and function; and 3): E. phagocytophila group ehrlichiae potentially influence host cell function by expressing proteins that directly interact with DNA regulatory components in the host cell's chromosomes. These hypotheses will be tested using five specific aims: i) demonstrate the morphology of the ehrlichia-host cell membrane interaction in HL60 cells in various differentiated states; ii) characterize the role of E. phagocytophila group MSPs as adhesins and the role of CD15-associated structures as the major surface receptors by which ehrlichiae attach and enter cells; iii) characterize the effects of ehrlichial binding and internalization on host granulocyte cell surface adhesion molecule expression, granulocyte-endothelial cell adherence, phagocytic activity, activation/deactivation, microbial killing, and cytokine/chemokine expression; and iv) identify the chromosomal ligand for the E. phagocytophila group ankyrin protein, EPANK1.