The Wnt signaling network, an ancient signaling system governing ontogeny and homeostatic processes, has recently been identified to exert immunoregulatory functions in a variety of inflammatory and infectious disease settings including tuberculosis. In this study, we show that Wnt6 is expressed in granulomatous lesions in the lung of Mycobacterium tuberculosis-infected mice. We identified foamy macrophage-like cells as the primary source of Wnt6 in the infected lung and uncovered a TLR-MyD88-NF-?B-dependent mode of induction in bone marrow-derived macrophages. Analysis of Wnt6-induced signal transduction revealed a pertussis toxin-sensitive, ERK-mediated, but ?-catenin-independent induction of c-Myc, a master regulator of cell proliferation. Increased Ki-67 mRNA expression levels and enhanced thymidine incorporation in Wnt6-treated macrophage cultures demonstrate a proliferation-promoting effect on murine macrophages. Further functional studies in M. tuberculosis-infected macrophages using Wnt6 conditioned medium and Wnt6-deficient macrophages uncovered a Wnt6-dependent induction of macrophage Arginase-1 and downregulation of TNF-?. This identifies Wnt6 as a novel factor driving macrophage polarization toward an M2-like phenotype. Taken together, these findings point to an unexpected role for Wnt6 in macrophage differentiation in the M. tuberculosis-infected lung.
In infection experiments with genetically distinct Mycobacterium tuberculosis complex (MTBC) strains, we identified clade-specific virulence patterns in human primary macrophages and in mice infected by the aerosol route, both reflecting relevant model systems. Exclusively human-adapted M. tuberculosis lineages, also termed clade I, comprising "modern" lineages, such as Beijing and Euro-American Haarlem strains, showed a significantly enhanced capability to grow compared to that of clade II strains, which include "ancient" lineages, such as, e.g., East African Indian or M. africanum strains. However, a simple correlation of inflammatory response profiles with strain virulence was not apparent. Overall, our data reveal three different pathogenic profiles: (i) strains of the Beijing lineage are characterized by low uptake, low cytokine induction, and a high replicative potential, (ii) strains of the Haarlem lineage by high uptake, high cytokine induction, and high growth rates, and (iii) EAI strains by low uptake, low cytokine induction, and a low replicative potential. Our findings have significant implications for our understanding of host-pathogen interaction and factors that modulate the outcomes of infections. Future studies addressing the underlying mechanisms and clinical implications need to take into account the diversity of both the pathogen and the host.
Bacterial infections are known to cause severe health-threatening conditions, including sepsis. All attempts to get this disease under control failed in the past, and especially in times of increasing antibiotic resistance, this leads to one of the most urgent medical challenges of our times. We designed a peptide to bind with high affinity to endotoxins, one of the most potent pathogenicity factors involved in triggering sepsis. The peptide Pep19-2.5 reveals high endotoxin neutralization efficiency in vitro, and here, we demonstrate its antiseptic/anti-inflammatory effects in vivo in the mouse models of endotoxemia, bacteremia, and cecal ligation and puncture, as well as in an ex vivo model of human tissue. Furthermore, we show that Pep19-2.5 can bind and neutralize not only endotoxins but also other bacterial pathogenicity factors, such as those from the Gram-positive bacterium Staphylococcus aureus. This broad neutralization efficiency and the additive action of the peptide with common antibiotics makes it an exceptionally appropriate drug candidate against bacterial sepsis and also offers multiple other medication opportunities.
Here we describe a novel approach for the isolation and biochemical characterization of pathogen-containing compartments from primary cells: We developed a lipid-based procedure to magnetically label the surface of bacteria and visualized the label by scanning and transmission electron microscopy (SEM, TEM). We performed infection experiments with magnetically labeled Mycobacterium avium, M. tuberculosis and Listeria monocytogenes and isolated magnetic bacteria-containing phagosomes using a strong magnetic field in a novel free-flow system. Magnetic labeling of M. tuberculosis did not affect the virulence characteristics of the bacteria during infection experiments addressing host cell activation, phagosome maturation delay and replication in macrophages in vitro. Biochemical analyses of the magnetic phagosome-containing fractions provided evidence of an enhanced presence of bacterial antigens and a differential distribution of proteins involved in the endocytic pathway over time as well as cytokine-dependent changes in the phagosomal protein composition. The newly developed method represents a useful approach to characterize and compare pathogen-containing compartments, in order to identify microbial and host cell targets for novel anti-infective strategies.
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