HOST GENE EXPRESSION DURING GRANULOCYTIC EHRLICHIOSIS

人类粒细胞性埃立克体病(HGE)是一种新发现的、由蜱类传播的感染。HGE制剂使中性粒细胞定植，导致发热、白细胞减少和血小板减少。HGE试剂可在早幼粒细胞HL-60细胞中增殖，有利于体外发病机制的研究。我们实验室的初步研究表明，在HL-60细胞停留期间，HGE通过下调NADPH氧化酶的组成部分gp91Phox的转录来防止呼吸爆发，机制尚不清楚。破译HGE制剂如何抑制gp91Phox的表达，并识别感染期间差异表达的其他宿主细胞基因，对于合理设计治疗和疫苗至关重要。因此，使用维甲酸诱导的HL-60细胞、偏粒系PLB-985细胞和粒细胞埃立克体病的小鼠模型进行转录研究，以确定HGE是否分别在分化的髓系细胞和哺乳动物感染中抑制gp91Phox的表达。RT-PCR和免疫印迹分析将评估HGE是否通过抑制gp91Phox转录调节因子的表达来阻止gp91Phox转录。电泳迁移率改变分析将确定gp91Phox转录调节因子的结合是否受到HGE试剂的抑制。这些研究除了提供对HGE的更多了解外，还可能为其他微生物病原体的细胞内生存机制提供线索。

Human granulocytic ehrlichiosis (HGE) is a newly recognized, tick-borne infection. The HGE agent colonizes neutrophils, resulting in fever, leukopenia, and thrombocytopenia. The HGE agent can be propagated in promyelocytic HL-60 cells, facilitating in vitro pathogenesis studies. Preliminary studies in our laboratory suggest that during residence in HL-60 cells, the HGE agent prevents the respiratory burst by downregulating transcription of gp91phox, an integral component of NADPH oxidase, by a currently undefined mechanism. Deciphering how the HGE agent inhibits gp91phox expression and identifying other host cell genes that are differentially expressed during infection are crucial to the rational design of therapies and vaccines. Therefore, transcriptional studies using retinoic acid-induced HL-60 cells, metamyelocytic PLB-985 cells, and a murine model of granulocytic ehrlichiosis will be performed to determine if gp91phox expression is inhibited by the HGE agent during residence in differentiated myeloid cells and mammalian infection, respectively. RT-PCR and immunoblot analyses will assess whether the HGE agent blocks gp91phox transcription by inhibiting expression of gp91phox transcriptional regulators. Electrophoretic mobility shift assays will define whether binding of the gp91phox transcriptional regulators is inhibited by the HGE agent. These studies, in addition to providing a greater understanding of HGE, may also offer clues as to the intracellular survival mechanisms of other microbial pathogens.