Impact of Antibiotics and PBP2a on the Immunopathology of MRSA pneumonia

描述(申请人提供)：耐甲氧西林金黄色葡萄球菌(MRSA)在全球范围内的迅速扩散对公众健康构成了重大威胁，并产生了已被证明比甲氧西林敏感金黄色葡萄球菌(MSSA)感染更难治疗的感染。原因尚不清楚，但普通抗生素有效性降低和长期炎症可能是感染的病理生理学原因，特别是在肺部。虽然MRSA可能对常见抗生素具有抗药性是不言而喻的，但增强或延长炎症的来源可能与细菌的持久性有关，也可能与细菌的持久性无关。金黄色葡萄球菌(像所有革兰氏阳性细菌一样)的细胞壁主要由高度交联的肽聚糖聚合物组成，我们实验室最近的工作表明，吞噬细胞降解金黄色葡萄球菌肽聚糖可以通过释放细菌DNA等促炎细胞内成分来强烈增强炎症反应。金黄色葡萄球菌主动修饰其肽聚糖，使其抵抗降解并抑制炎症反应，当这一作用被抑制时，金黄色葡萄球菌在体外诱导巨噬细胞产生更多的细胞因子，并在小鼠中引起更多的免疫病理。我们进一步发现，抗生素诱导的金黄色葡萄球菌ALS细胞壁肽聚糖合成的亚致死性改变导致巨噬细胞引发更强大的炎症反应。耐甲氧西林金黄色葡萄球菌通过诱导PBP2a的表达而产生抗药性，PBP2a是一种对β-内酰胺类抗生素具有抗药性的肽聚糖合成酶。然而，据报道，PBP2a的表达会导致肽聚糖的合成和结构发生变化，我们现在观察到，这直接增强了巨噬细胞中的炎症信号。基于这些发现，我们得出了一个令人惊讶的假设，即特定的抗生素实际上可能会导致免疫病理学在MRSA感染期间变得更加明显。我们将在两个目标上检验这一假设。在第一个目标中，我们将直接检测PBP2a和临床上常用的抗生素对金黄色葡萄球菌体外炎症能力的影响。利用基因敲除小鼠的细胞，我们将确定炎症能力变化的潜在机制。在第二个目标中，我们将转移到活体小鼠肺炎模型，以确定抗生素和MRSA表达PBP2a对免疫病理的影响。 公共卫生相关性：与甲氧西林敏感金黄色葡萄球菌(MSSA)相比，耐甲氧西林金黄色葡萄球菌(MRSA)引起的肺部感染似乎更严重，更难治疗。在这里，我们研究了细胞壁活性抗生素和编码甲氧西林耐药性的基因都有助于增强免疫病理学的假设。如果我们的假设是正确的，我们的研究可能会发现MRSA肺炎患者发病的一个重要原因，并可能通过解决炎症造成的损害来为治疗MRSA感染的更有效方法铺平道路。

DESCRIPTION (provided by applicant): The rapid worldwide proliferation of Methicillin-Resistant S. aureus (MRSA) poses a major threat to public health and produces infections that have proven to be more difficult to treat compared to Methicillin- Susceptible S. aureus (MSSA) infections. The reason for this is unknown, but reduced effectiveness of common antibiotics and prolonged inflammation likely contribute to the pathophysiology of infection, especially in the lung. While it is self-evident that MRSA may be resistant to common antibiotics, the source of enhanced or prolonged inflammation may or may not be related to perseverance of the bacteria. The cell wall of S. aureus (like all Gram-positive bacteria) is made up primarily of highly cross-linked peptidoglycan polymer, and recent work from our labs has suggested that degradation of the S. aureus peptidoglycan by phagocytes strongly enhances inflammatory responses by releasing pro-inflammatory intracellular components such as bacterial DNA. S. aureus actively modifies its peptidoglycan to make it resistant to degradation and suppresses inflammatory responses, and when this effect is inhibited, S. aureus induces greater cytokine production from macrophages in vitro and causes more immunopathology in mice. We have further found that sub-lethal alterations in cell wall peptidoglycan synthesis induced by antibiotics in S. aureus als causes macrophages to elicit a more powerful inflammatory response. MRSA becomes antibiotic-resistant through induced expression of PBP2A, a peptidoglycan synthesizing enzyme that is resistant to ?-lactam antibiotics. Expression of PBP2A, however, has been reported to cause alterations in peptidoglycan synthesis and structure, and we have now observed that this directly enhances inflammatory signaling in macrophages. Based on these findings, we come to the surprising hypothesis that specific antibiotics may actually cause immunopathology to become more pronounced during MRSA infection. We will test this hypothesis in two aims. In the first aim, we will examine directly the effects of PBP2A and antibiotics typically used in the clinic on the inflammatory capacity of S. aureus in vitro. Using cells from knockout mice we will identify the mechanisms underlying alterations in inflammatory capacity. In the second aim, we will move to in vivo mouse pneumonia models to determine the effects of antibiotics and PBP2A expression by MRSA on immunopathology. PUBLIC HEALTH RELEVANCE: Compared to Methicillin-Sensitive S. aureus (MSSA), Methicillin-Resistant S. aureus (MRSA) causes lung infections that appear more severe and are harder to treat. Here we investigate the hypothesis that cell wall active antibiotics and the gene encoding methicillin resistance both contribute to increased immunopathology. If our hypothesis is correct, our research could uncover an important cause of morbidity in MRSA pneumonia patients, and could pave the way for more effective ways to treat MRSA infections by addressing the damage caused by inflammation.