Neutrophils Protect Against Staphylococcus aureus Endocarditis Progression Independent of Extracellular Trap Release

Abstract

Background: Infective endocarditis (IE) is characterized by an infected thrombus at the heart valves. How bacteria bypass the immune system and cause these thrombi remains unclear. Neutrophils releasing NETs (neutrophil extracellular traps) lie at this interface between host defense and coagulation. We aimed to determine the role of NETs in IE immunothrombosis. Methods: We used a murine model of Staphylococcus aureus endocarditis in which IE is provoked on inflamed heart valves and characterized IE thrombus content by immunostaining identifying NETs. Antibody-mediated neutrophil depletion and neutrophil-selective PAD4 (peptidylarginine deiminase 4)-knockout mice were used to clarify the role of neutrophils and NETs, respectively. S. aureus mutants deficient in key virulence factors related to immunothrombosis (nucleases or staphylocoagulases) were investigated. Results: Neutrophils releasing NETs were present in infected thrombi and within cellular infiltrates in the surrounding vasculature. Neutrophil depletion increased occurrence of IE, whereas neutrophil-selective impairment of NET formation did not alter IE occurrence. Absence of S. aureus nuclease, which degrades NETs, did not affect endocarditis outcome. In contrast, absence of staphylocoagulases (coagulase and von Willebrand factor binding protein) led to improved survival, decreased bacteremia, smaller infiltrates, and decreased tissue destruction. Significantly more NETs were present in these vegetations, which correlated with decreased bacteria and cell death in the adjacent vascular wall. Conclusions: Neutrophils protect against IE independent of NET release. Absence of S. aureus coagulases, but not nucleases, reduced IE severity and increased NET levels. Staphylocoagulase-induced fibrin likely hampers NETs from constraining infection and the resultant tissue damage, a hallmark of valve destruction in IE.

Neutrophil-driven cardiac damage during invasiveStreptococcus pneumoniaeinfection is regulated by CD73

“…This bacterium however, is known to produce C5a peptidase and CspA serine protease that impair or blunt PMN recruitment and response at the site of infection [51]. In S. aureus -mediated mouse endocarditis model, infiltration of PMNs in infected thrombi is shown to be important to limit the disease progression [52]. However, this influx of leukocytes around the foci of infection was also accompanied by extensive proteolytic cardiac tissue damage and cellular apoptosis [52].…”

“…In S. aureus -mediated mouse endocarditis model, infiltration of PMNs in infected thrombi is shown to be important to limit the disease progression [52]. However, this influx of leukocytes around the foci of infection was also accompanied by extensive proteolytic cardiac tissue damage and cellular apoptosis [52]. On the other hand, during infective endocarditis caused by E. faecalis , release of metalloproteinases and elastase by PMNs infiltrating in the cardiac tissue were suggested to promote the formation and growth of bacteria containing thrombi in the heart, and induce myocardial damage [53].…”

The coming of age of neutrophil extracellular traps in thrombosis: Where are we now and where are we headed?

“…We recently reported that NETs are not sufficient to mitigate S. aureus-mediated IE, whereas neutrophil depletion showed the importance of neutrophils in reducing the incidence of IE. 176 This is due to S. aureus's staphylocoagulases that likely form a fibrin shield that keeps the bacteria away from neutrophils; therefore, their potential impact on bacteria is circumvented by this physical barrier. In the absence of these coagulases, NETs play an important role in preventing spread of the bacteria into the area outside of the vessel wall.…”

Peptidylarginine deiminase 4 and ADAMTS13 activity in Staphylococcus aureus bacteraemia

“…This high mortality, the rising incidence of S. aureus infections and the growing antibiotic resistance emphasize the need to address this infectious disease as a global health priority [5]. During infection, S. aureus triggers neutrophil activation and induces the release of neutrophil extracellular traps (NETs), or extracellular DNA strands covered with histones and other proteins that are released from activated neutrophils [6][7][8] (figure 1). Peptidylarginine deiminase 4 (PAD4) is a citrullinating enzyme that contributes to NET formation by converting specific arginine residues on histones to citrulline, which results in the loss of positive charges and subsequent dissociation of chromatin fibres and NET release [9].…”

“…NETs play a direct role in antimicrobial defence mechanisms by trapping and thus sequestering bacteria [6,7,14]. Additionally, NETs interact with plasma proteins such as von Willebrand factor (VWF) [15,16].…”