Cardiac microlesion formation during invasive pneumococcal disease

描述（由申请人提供）：大约1/5因社区获得性肺炎（CAP）住院的成年人经历心律失常、充血性心力衰竭和/或心肌梗死，这是导致死亡率的主要原因。我们发现，作为CAP的主要病因，肺炎链球菌可在侵袭性肺炎球菌病（IPD）期间进入心肌，形成充满细胞外肺炎球菌的显微空泡性病变（即微性病变）。在患有IPD的小鼠中，心脏微病变的形成伴随着异常的心脏电生理，表明持续的心力衰竭。与其他细菌引起的心脏感染形成鲜明对比的是，肺炎链球菌心脏微病变的形成是显著的，因为完全没有浸润的免疫细胞。发生这种情况的原因尚不清楚，但缺乏宿主反应无疑会导致心脏损伤。我们假设肺炎链球菌在心脏微病变内形成免疫静止生物膜。我们的初步结果支持这一观点，并证明生物膜肺炎链球菌（BF-Spn）不会激发宿主细胞的NFκB激活，也不会导致促炎细胞因子的产生。我们的目标是辨别

DESCRIPTION (provided by applicant): Approximately 1/5 of adults hospitalized for community-acquired pneumonia (CAP) experience cardiac arrhythmia, congestive heart failure, and/or myocardial infarction and this contributes substantially to mortality. We have made the novel and seminal observation that Streptococcus pneumoniae, the leading cause of CAP, can enter the myocardium during invasive pneumococcal disease (IPD) and forms microscopic vacuolar lesions (i.e. microlesions) filled with extracellular pneumococci. In mice with IPD, cardiac microlesion formation was concomitant with aberrant cardiac electrophysiology indicative of ongoing heart failure. In stark contrast to cardiac infections caused by other bacteria, S. pneumoniae cardiac microlesion formation is remarkable due to the complete absence of infiltrated immune cells. Why this occurs is unknown, yet the lack of a host response is undoubtedly permissive for heart damage. We hypothesize that S. pneumoniae forms an immunoquiescent biofilm within the cardiac microlesion. Our preliminary results support this notion and demonstrate that biofilm S. pneumoniae (BF-Spn) does not incite NFκB activation from host cells nor results in the production of pro-inflammatory cytokines. Our goal is to discern the host-pathogen interactions that occur during cardiac microlesion formation and identify targets for intervention. Aim 1: Determine the requirement for biofilm formation during pneumococcal cardiac microlesion formation. We will examine cardiac microlesions for the presence of biofilm extracellular matrix components and test isolated S. pneumoniae for the biofilm phenotype. We will test the ability of S. pneumoniae mutants deficient in the ability to form biofilms in vitro for their ability to form cardiac microlesions in vivo. We will compare the cytokine and chemokine response of cardiomyocytes exposed to planktonic (P-Spn) and BF-Spn. This aim will directly test if S. pneumoniae within cardiac microlesions are in an immunoquiescent biofilm. Aim 2: Characterize S. pneumoniae within cardiac microlesions and identify novel virulence determinants required for their formation. Using RNA-sequencing we will characterize and then compare the gene expression profile of S. pneumoniae in cardiac microlesions to those in the blood of infected mice. Genes with high in vivo gene expression will be deleted and isogenic mutants tested for the ability to cause cardiac microlesions. This aim will characterize how S. pneumoniae adapts to growth within the heart and identify in an unbiased manner targets for intervention. Aim 3: Characterize the cardiomyocyte response to pneumolysin exposure. We will measure the cytokine/chemokine response and susceptibility of cardiomyocytes to pneumolysin, the S. pneumoniae pore forming toxin, and bacterial cell wall. We will test if S. pneumoniae activates the NLRP3 inflammasome within cardiomyocytes. We will determine if sub-lytic levels of pneumolysin trigger the intrinsic pathway of apoptosis in cardiomyocytes. This aim will discern if cardiomyocytes exposed to S. pneumoniae undergo a cell death program that is not inflammatory.