24 research outputs found
Pneumolysin binds to the mannose receptor C type 1 (MRC-1) leading to anti-inflammatory responses and enhanced pneumococcal survival
Streptococcus pneumoniae (the pneumococcus) is a major cause of mortality and morbidity globally, and the leading cause of death in children under 5 years old. The pneumococcal cytolysin pneumolysin (PLY) is a major virulence determinant known to induce pore-dependent pro-inflammatory responses. These inflammatory responses are driven by PLY–host cell membrane cholesterol interactions, but binding to a host cell receptor has not been previously demonstrated. Here, we discovered a receptor for PLY, whereby pro-inflammatory cytokine responses and Toll-like receptor signalling are inhibited following PLY binding to the mannose receptor C type 1 (MRC-1) in human dendritic cells and mouse alveolar macrophages. The cytokine suppressor SOCS1 is also upregulated. Moreover, PLY–MRC-1 interactions mediate pneumococcal internalization into non-lysosomal compartments and polarize naive T cells into an interferon-γlow, interleukin-4high and FoxP3+ immunoregulatory phenotype. In mice, PLY-expressing pneumococci colocalize with MRC-1 in alveolar macrophages, induce lower pro-inflammatory cytokine responses and reduce neutrophil infiltration compared with a PLY mutant. In vivo, reduced bacterial loads occur in the airways of MRC-1-deficient mice and in mice in which MRC-1 is inhibited using blocking antibodies. In conclusion, we show that pneumococci use PLY–MRC-1 interactions to downregulate inflammation and enhance bacterial survival in the airways. These findings have important implications for future vaccine design
Circulating Pneumolysin Is a Potent Inducer of Cardiac Injury during Pneumococcal Infection
Streptococcus pneumoniae accounts for more deaths worldwide than any other single pathogen through diverse disease manifestations including pneumonia, sepsis and meningitis. Life-threatening acute cardiac complications are more common in pneumococcal infection compared to other bacterial infections. Distinctively, these arise despite effective antibiotic therapy. Here, we describe a novel mechanism of myocardial injury, which is triggered and sustained by circulating pneumolysin (PLY). Using a mouse model of invasive pneumococcal disease (IPD), we demonstrate that wild type PLY-expressing pneumococci but not PLY-deficient mutants induced elevation of circulating cardiac troponins (cTns), well-recognized biomarkers of cardiac injury. Furthermore, elevated cTn levels linearly correlated with pneumococcal blood counts (r=0.688, p=0.001) and levels were significantly higher in non-surviving than in surviving mice. These cTn levels were significantly reduced by administration of PLY-sequestering liposomes. Intravenous injection of purified PLY, but not a non-pore forming mutant (PdB), induced substantial increase in cardiac troponins to suggest that the pore-forming activity of circulating PLY is essential for myocardial injury in vivo. Purified PLY and PLY-expressing pneumococci also caused myocardial inflammatory changes but apoptosis was not detected. Exposure of cultured cardiomyocytes to PLY-expressing pneumococci caused dose-dependent cardiomyocyte contractile dysfunction and death, which was exacerbated by further PLY release following antibiotic treatment. We found that high PLY doses induced extensive cardiomyocyte lysis, but more interestingly, sub-lytic PLY concentrations triggered profound calcium influx and overload with subsequent membrane depolarization and progressive reduction in intracellular calcium transient amplitude, a key determinant of contractile force. This was coupled to activation of signalling pathways commonly associated with cardiac dysfunction in clinical and experimental sepsis and ultimately resulted in depressed cardiomyocyte contractile performance along with rhythm disturbance. Our study proposes a detailed molecular mechanism of pneumococcal toxin-induced cardiac injury and highlights the major translational potential of targeting circulating PLY to protect against cardiac complications during pneumococcal infections
Service provider success: story of Liberty Autos
The elements for success in the service industry for many organizations have been studied and quantified. They range from effective performance evaluation and management to adequately implementing seven P\u27s. However, the success story of every organization is unique. What might work for one organization might not for another as every company is inherently distinctive. In this paper, the strategic framework behind the success of Liberty Automobiles Dubai is captured. The primary data necessary for this paper was collected through in-depth interviews with the managers for sales, service and marketing as well as the group general manager. Subsequently, the key areas that led to the success of Liberty Autos will be discussed in this paper
Brand personification as a service marketing mechanism
Today the business environment is becoming increasingly competitive, with a vast array of products that can no longer be differentiated on product quality or features. As such, many businesses are trying to differentiate themselves from their competitors using their provision of service. One concept that could possibly enhance a firm\u27s provision of service is brand personification. Accordingly, this paper, will discuss how Liberty Autos, a successful automobile retailer in the UAE used \u27brand personification\u27 as a services marketing mechanism
The Pneumococcal Polysaccharide Capsule and Pneumolysin Differentially Affect CXCL8 and IL-6 Release from Cells of the Upper and Lower Respiratory Tract
The polysaccharide capsule and pneumolysin toxin are major virulence factors of the human bacterial pathogen Streptococcus pneumoniae. Colonization of the nasopharynx is asymptomatic but invasion of the lungs can result in invasive pneumonia. Here we show that the capsule suppresses the release of the pro-inflammatory cytokines CXCL8 (IL-8) and IL-6 from the human pharyngeal epithelial cell line Detroit 562. Release of both cytokines was much less from human bronchial epithelial cells (iHBEC) but levels were also affected by capsule. Pneumolysin stimulates CXCL8 release from both cell lines. Suppression of CXCL8 homologue (CXCL2/MIP-2) release by the capsule was also observed in vivo during intranasal colonization of mice but was only discernable in the absence of pneumolysin. When pneumococci were administered intranasally to mice in a model of long term, stable nasopharyngeal carriage, encapsulated S. pneumoniae remained in the nasopharynx whereas the nonencapsulated pneumococci disseminated into the lungs. Pneumococcal capsule plays a role not only in protection from phagocytosis but also in modulation of the pro-inflammatory immune response in the respiratory tract
Neutrophil IL-1β Processing Induced by Pneumolysin Is Mediated by the NLRP3/ASC Inflammasome and Caspase-1 Activation and Is Dependent on K+ Efflux
Although neutrophils are the most abundant cells in acute infection and inflammation, relatively little attention has been paid to their role in inflammasome formation and IL-1β processing. In the present study, we investigated the mechanism by which neutrophils process IL-1β in response to Streptococcus pneumoniae. Using a murine model of S. pneumoniae corneal infection, we demonstrated a requirement for IL-1β in bacterial clearance, and we showed that Nod-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC), and caspase-1 are essential for IL-1β production and bacterial killing in the cornea. Neutrophils in infected corneas had multiple specks with enzymatically active caspase-1 (YVAD-FLICA 660), and bone marrow neutrophils stimulated with heat-killed S. pneumoniae (signal 1) and pneumolysin (signal 2) exhibited multiple specks when stained for NLRP3, ASC, or Caspase-1. High-molecular mass ASC complexes were also detected, consistent with oligomer formation. Pneumolysin induced K(+) efflux in neutrophils, and blocking K(+) efflux inhibited caspase-1 activation and IL-1β processing; however, neutrophils did not undergo pyroptosis, indicating that K(+) efflux and IL-1β processing is not a consequence of cell death. There was also no role for lysosomal destabilization or neutrophil elastase in pneumolysin-mediated IL-1β processing in neutrophils. Taken together, these findings demonstrate an essential role for neutrophil-derived IL-1β in S. pneumoniae infection, and they elucidate the role of the NLRP3 inflammasome in cleavage and secretion of IL-1β in neutrophils. Given the ubiquitous presence of neutrophils in acute bacterial and fungal infections, these findings will have implications for other microbial diseases
Effect of capsule and pneumolysin on CXCL8 and IL-6 induction in human nasopharyngeal and bronchial epithelial cells.
<p>Detroit 562 nasopharyngeal epithelial cells (A and B) and bronchial epithelial cells (C and D) were assessed for CXCL8 (A and C) and IL-6 (B and D) release after exposure to wild type or mutant pneumococcal strains. All experiments were performed in triplicate at each of three CFU concentrations (1, 1.5 and 2 × 10<sup>6</sup>) and the results pooled for each strain. Note different scales of Y axes. Error bars indicate SEM. * indicates significant difference.</p
Capsule did not affect colonization of the nasopharynx but only nonencapsulated strains reached the lungs.
<p>Each symbol represents the CFU from the nasopharynx or lungs of an individual mouse on days 1, 3, 8 and 15 after intranasal inoculation. (No bacteria were detected at day 0 before any bacteria were administered.) Horizontal bars indicate means.</p
Effect of capsule and pneumolysin on CXCL8 homologue induction in the mouse nasopharynx.
<p>CXCL8 homologue (CXCL2/MIP-2) detected in nasopharyngeal homogenate of mice three days after exposure to wild type or mutant pneumococci expressed as a percentage of the value obtained with the wild type strain. Error bars indicate SEM. * indicates significant difference from value of the parent strain.</p
PLY at sub-lytic doses adversely affect cardiomyocyte function <i>in vitro</i>.
<p>(<b>A</b>) Viability of HL-1 cells was assessed 30 min after incubation with increasing concentrations of PLY and PdB using the WST-8 assay. Viability of untreated cells (UT) was set at 100%. Data are presented as Mean±SD. * <i>p</i><0.05 (n = 4). (<b>B</b>) Time course of HL-1 cell viability after incubation with 1.5 μg/ml PLY or PdB. *<i>p</i><0.05 (n = 3). (<b>C</b>) Effects of increasing concentrations of PLY on the total number of spontaneously contracting HL-1 cells over time. Data are presented as Mean±SD. *<i>p</i><0.05 (n = 4). (<b>D</b>) Representative traces of cardiomyocyte contraction before and after PLY and PdB treatment (n = 4). (<b>E-I</b>) Effects of PLY and PdB (1.0 μg/ml) on Peak Shortening (E), +dL/dt (F), TTP (G), tR<sub>90</sub> (H) and -dL/dt (I) of HL-1 cells after 30 min treatment are presented as Mean±SD. *<i>p</i><0.05 (n = 9 from 3 independent experiments).</p