Microbial Response to Neutrophil Phagocytosis

描述（由申请人提供）：中性粒细胞是宿主防御细菌和真菌病原体的关键组成部分。当中性粒细胞在数量或功能上严重减少时，宿主对严重感染变得高度敏感。中性粒细胞通常在短时间内吸收并杀死细菌。为了更好地了解细菌对吞噬的反应，基因组阵列将用于评估被中性粒细胞摄入的大肠杆菌细胞mRNA丰度的变化。我们希望了解摄入的细菌如何优先响应吞噬细胞中产生的压力，以及这些反应是否能提高微生物的存活率。由吞噬细胞NADPH氧化酶（CGD，慢性肉芽肿病）或髓过氧化物酶介导的抗菌系统中中性粒细胞缺陷的影响将与正常中性粒细胞进行比较。最初的研究表明，正常的中性粒细胞，而不是CGD中性粒细胞，会引起由细菌转录因子OxyR介导的抗氧化反应。氧r基因的破坏产生了一种细菌菌株，这种菌株被正常中性粒细胞杀死的几率是正常中性粒细胞的10倍。在升肾盂肾炎小鼠模型中，oxyR菌株的毒力也明显减弱。本研究旨在更全面地表征氧合受体的作用，并探索其他主要转录反应对中性粒细胞吞噬和分离抗菌模型系统的影响。它还寻求实施一种方法，强调检测致病性大肠杆菌菌株中存在的吞噬诱导的mRNA转录物变化，而不是在实验室适应的菌株中。后一个目标的驱动假设是：病原体特异性转录物对致病表型有重大贡献。被吞噬细菌的表达谱似乎提供了关于吞噬液泡条件的有用信息，以及对这种敌对环境作出反应的重要细菌防御。

DESCRIPTION (provided by applicant): Neutrophils are a key component of host defense against bacterial and fungal pathogens. When neutrophils are severely diminished in number or function, the host becomes highly susceptible to serious infection. Neutrophils typically ingest and kill bacteria within a short space of time. In order to gain a better understanding of the bacterial response to phagocytosis, genomic arrays will be used to assess changes in mRNA abundance for Escherichia coli cells ingested by neutrophils. We expect to learn how ingested bacteria prioritize responses to stresses generated in the phagocyte and whether these responses enhance microbial survival. The effects of neutrophils deficient in antimicrobial systems mediated by the phagocyte NADPH oxidase (CGD, chronic granulomatous disease) or the enzyme myeloperoxidase will be compared with normal neutrophils. Initial studies indicate that normal neutrophils, but not CGD neutrophils, evoke an anti-oxidant response mediated by the bacterial transcription factor, OxyR. Disruption of the oxyR gene generated a bacterial strain that was 10-fold more susceptible to killing by normal neutrophils. The oxyR strain also appears to be markedly attenuated for virulence in a mouse model of ascending pyelonephritis. This proposal seeks to characterize oxyR effects more fully, and to explore the effects of selected other major transcriptional responses both to neutrophil phagocytosis and to isolated antimicrobial model systems. It also seeks to implement methodology that will emphasize detection of phagocytosis-induced changes for mRNA transcripts present in a pathogenic E. coli strain but not in a laboratory-adapted strain. The driving hypothesis for the latter goal being that: pathogen-specific transcripts contribute substantially to the pathogenic phenotype. Expression profiling of phagocytosed bacteria appears to provide useful information about conditions in the phagocytic vacuole and about important bacterial defenses mounted in response to this hostile environment.