Goldmann, Oliver; von Köckritz-Blickwede, Maren; Höltje, Claudia; Chhatwal, Gursharan S; Geffers, Robert; Medina, Eva (2007-08)
The complex response of murine macrophages to infection with Streptococcus pyogenes was investigated at the level of gene expression with a high-density oligomer microarray. More than 400 genes were identified as being differentially regulated. Many of the up-regulated genes encode molecules involved in the immune response and in inflammation, transcription, signaling, apoptosis, the cell cycle, electron transport, and cell adhesion. Of particular interest was the up-regulation of proinflammatory cytokines, typical of the classically activated macrophages (M1 phenotype), such as tumor necrosis factor alpha, interleukin 1 (IL-1), and IL-6, and as well as the up-regulation of anti-inflammatory mediators, such as IL-1 decoy receptor and IL-10, associated with alternative macrophage activation (M2 phenotype). Furthermore, the gene encoding inducible nitric oxide synthase (iNOS), an enzyme typically implicated in classical activation, was not induced in infected macrophages. Instead, the gene encoding arginase, a competitor for the iNOS substrate arginine involved in the alternative activation pathway, was up-regulated in S. pyogenes-infected cells. Thus, the microarray-based gene expression analysis demonstrated that S. pyogenes induces an atypical activation program in macrophages, with some but not all features of the classical or alternative activation phenotypes. The microarray data also suggested that the bactericidal activity of macrophages against S. pyogenes is mediated by phagocyte oxidase, as p47phox was up-regulated in infected cells. Indeed, the in vivo and in vitro killing of S. pyogenes was markedly diminished in the absence of functional phagocyte (p47(phox-/-)) but not in the absence of iNOS (iNOS(-/-)). An understanding of how macrophages respond to S. pyogenes at the molecular level may facilitate the development of new therapeutic paradigms.
Staphylococcus aureus is a frequent cause for serious, chronic and therapy-refractive infections in spite of susceptibility to antibiotics in vitro. In chronic infections, altered bacterial phenotypes, such as small colony variants (SCVs), have been found. Yet, it is largely unclear whether the ability to interconvert from the wild-type to the SCV phenotype is only a rare clinical and/or just laboratory phenomenon or is essential to sustain an infection. Here, we performed different long-term in vitro and in vivo infection models with S. aureus and we show that viable bacteria can persist within host cells and/or tissues for several weeks. Persistence induced bacterial phenotypic diversity, including SCV phenotypes, accompanied by changes in virulence factor expression and auxotrophism. However, the recovered SCV phenotypes were highly dynamic and rapidly reverted to the fully virulent wild-type form when leaving the intracellular location and infecting new cells. Our findings demonstrate that bacterial phenotype switching is an integral part of the infection process that enables the bacteria to hide inside host cells, which can be a reservoir for chronic and therapy-refractive infections.
Goldmann, Oliver; Lehne, Sabine; Medina, Eva (2010-04)
Epidemiological studies have shown that the elderly are at higher risk of severe Streptococcus pyogenes infections. In this study, we used a mouse model that displays the age-related loss of resistance to S. pyogenes infection seen in humans to investigate the impaired immune mechanism underlying the age-associated susceptibility to this pathogen. Young (2-3 months old) and aged (>20 months old) BALB/c mice were subcutaneously or intravenously inoculated with S. pyogenes and their capacity to control infection was compared. Aged mice showed faster progression of disease, earlier morbidity, and increased mortality when compared with young animals. Since macrophages are critical for host defence against S. pyogenes, we investigated whether susceptibility of aged mice may be due to an age-associated decline in the functionality of these cells. Our results showed that macrophages from aged mice were as capable as those from young animals to uptake and kill S. pyogenes, but the number of resident tissue macrophages was significantly reduced in the aged host. Treatment of aged mice with macrophage colony-stimulating factor (M-CSF) significantly increased the number of resident macrophages and improved their response to infection. Our results indicate that treatment with M-CSF can restore, at least in part, the mechanisms affected by immunosenescence and enhance the natural resistance of aged mice to infection with S. pyogenes.
Loof, Torsten G; Deicke, Christin; Medina, Eva (2014)
The hemostatic system comprises platelet aggregation, coagulation and fibrinolysis and is a host defense mechanism that protects the integrity of the vascular system after tissue injury. During bacterial infections, the coagulation system cooperates with the inflammatory system to eliminate the invading pathogens. However, pathogenic bacteria have frequently evolved mechanisms to exploit the hemostatic system components for their own benefit. Streptococcus pyogenes, also known as Group A Streptococcus, provides a remarkable example of the extraordinary capacity of pathogens to exploit the host hemostatic system to support microbial survival and dissemination. The coagulation cascade comprises the contact system (also known as the intrinsic pathway) and the tissue factor pathway (also known as the extrinsic pathway), both leading to fibrin formation. During the early phase of S. pyogenes infection, the activation of the contact system eventually leads to bacterial entrapment within a fibrin clot, where S. pyogenes is immobilized and killed. However, entrapped S. pyogenes can circumvent the antimicrobial effect of the clot by sequestering host plasminogen on the bacterial cell surface that, after conversion into its active proteolytic form, plasmin, degrades the fibrin network and facilitates the liberation of S. pyogenes from the clot. Furthermore, the surface-localized fibrinolytic activity also cleaves a variety of extracellular matrix proteins, thereby enabling S. pyogenes to migrate across barriers and disseminate within the host. This review summarizes the knowledge gained during the last two decades on the role of coagulation/fibrinolysis in host defense against S. pyogenes as well as the strategies developed by this pathogen to evade and exploit these host mechanisms for its own benefit.
Infections by are a major cause of morbidity and mortality worldwide, often causing community-acquired pneumonia, otitis media and also bacteremia and meningitis. Studies on are mainly focused on its virulence or capacity to evade the host immune system, but little is known about the injury caused in lungs during a pneumococcal infection. Herein we investigated this issue comparing the proteome profile of lungs from infected mice with control mice by means of difference gel electrophoresis (DIGE) technology. In order to obtain reliable results three biological replicas were used, and four technical replicas were carried out in each biological replica. Proteomic comparison was performed at two time points: 24 and 48 h post infection. A total of 91 proteins were identified with different abundance. We found important changes in the protein profiles during pneumococcal infection mainly associated with regulation of vesicle-mediated transport, wound healing, and cytoskeleton organization. In conclusion, the results obtained show that the cytoskeleton of the host cell is modified in infection.
Upon the onset of inflammatory responses, bacterial pathogens are confronted with altered tissue microenvironments which can critically impact on their metabolic activity and growth. Changes in these parameters have however remained difficult to analyze over time, which would be critical to dissect the interplay between the host immune response and pathogen physiology. Here, we established an in vivo biosensor for measuring the growth rates of Staphylococcus aureus (S. aureus) on a single cell-level over days in an ongoing cutaneous infection. Using intravital 2-photon imaging and quantitative fluorescence microscopy, we show that upon neutrophil recruitment to the infection site and bacterial uptake, non-lethal dampening of S. aureus proliferation occurred. This inhibition was supported by NADPH oxidase activity. Therefore, reactive oxygen production contributes to pathogen containment within neutrophils not only by killing S. aureus, but also by restricting the growth rate of the bacterium.
After initial infection, the immune response that serves to restrict the invading pathogen needs to be tightly calibrated in order to avoid collateral immunopathological damage. This calibration is performed by specialized suppressor mechanisms, which are capable of dampening overwhelming or unremitting inflammation in order to prevent tissue damage. Myeloid-derived suppressor cells (MDSC) are emerging as key players in counter-balancing inflammatory responses and pathogenesis during infection. However, some pathogens are able to exploit the suppressive activities of MDSC to favor pathogen persistence and chronic infections. In this article, we review the current knowledge about the importance of MDSC in the context of bacterial, virus, parasites, and fungal infections.
Siegert, Jeannette; Sastalla, Inka; Chhatwal, Gursharan Singh; Medina, Eva (2006-02-01)
There is substantial evidence that host genetic factors are important in determining susceptibility to infection with group A streptococci (GAS). Several studies have revealed that, similarly to humans, a genetic component may be important in determining susceptibility to GAS infection in mice. Thus, C3H/HeN mice are much more susceptible to streptococcal infection than BALB/c mice. We have determined here whether vaccination makes genetically susceptible mice as capable as genetically resistant mice to control GAS infection. Resistant BALB/c and susceptible C3H/HeN mice were immunized either systemically with heat-killed GAS or through the mucosal route with an M protein-based subunit vaccine, and challenged with live bacteria. Vaccination elicited in both mouse strains similar levels of bactericidal anti-GAS IgG antibodies and also antigen-specific mucosal IgA. Vaccination provided mice of both strains with an increased and equal capacity to express immunity against GAS as indicated by the reduced level of bacteria in the organs and the ability of vaccinated mice to survive infection. Protection in vaccinated mice was dependent on the presence of T cell-dependent bactericidal antibodies as shown by the ability of serum elicited in immunocompetent mice but not of serum elicited in T cell-deficient nu/nu mice to passively transfer anti-GAS immunity. In conclusion, the results presented here demonstrated that the presence of anti-GAS specific, T cell-dependent bactericidal antibodies elicited after vaccination overcomes the innate genetic susceptibility of C3H/HeN mice and makes both resistant and susceptible mice equally capable of controlling GAS infection.
The release of extracellular traps (ETs) is a recently described mechanism of innate immune response to infection. Although ETs have been intensely investigated in the context of neutrophil antimicrobial effector mechanisms, other immune cells such as mast cells, eosinophils, and macrophages can also release these structures. The different ETs have several features in common, regardless of the type of cells from which they originated, including a DNA backbone with embedded antimicrobial peptides, proteases, and histones. However, they also exhibit remarkable individual differences such as the type of sub-cellular compartments from where the DNA backbone originates (e.g., nucleus or mitochondria), the proportion of responding cells within the pool, and/or the molecular mechanism/s underlying the ETs formation. This review summarizes the knowledge accumulated in recent years regarding the complex and expanding world of ETs and their role in immune function with particular emphasis on the role of other immune cells rather than on neutrophils exclusively.
Streptococcus pneumoniae is a leading cause of bacterial pneumonia worldwide. Given the critical role of dendritic cells (DCs) in regulating and modulating the immune response to pathogens, we investigated here the role of DCs in S. pneumoniae lung infections. Using a well-established transgenic mouse line which allows the conditional transient depletion of DCs, we showed that ablation of DCs resulted in enhanced resistance to intranasal challenge with S. pneumoniae. DCs-depleted mice exhibited delayed bacterial systemic dissemination, significantly reduced bacterial loads in the infected organs and lower levels of serum inflammatory mediators than non-depleted animals. The increased resistance of DCs-depleted mice to S. pneumoniae was associated with a better capacity to restrict pneumococci extrapulmonary dissemination. Furthermore, we demonstrated that S. pneumoniae disseminated from the lungs into the regional lymph nodes in a cell-independent manner and that this direct way of dissemination was much more efficient in the presence of DCs. We also provide evidence that S. pneumoniae induces expression and activation of matrix metalloproteinase-9 (MMP-9) in cultured bone marrow-derived DCs. MMP-9 is a protease involved in the breakdown of extracellular matrix proteins and is critical for DC trafficking across extracellular matrix and basement membranes during the migration from the periphery to the lymph nodes. MMP-9 was also significantly up-regulated in the lungs of mice after intranasal infection with S. pneumoniae. Notably, the expression levels of MMP-9 in the infected lungs were significantly decreased after depletion of DCs suggesting the involvement of DCs in MMP-9 production during pneumococcal pneumonia. Thus, we propose that S. pneumoniae can exploit the DC-derived proteolysis to open tissue barriers thereby facilitating its own dissemination from the local site of infection.
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