Early Host Defense Mechanism Research Articles

Factor H-related protein 1 promotes complement-mediated opsonization of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an important human opportunistic pathogen responsible for a wide range of infections. The complement system is the main early host defense mechanism to control these infections. P. aeruginosa counteracts complement attack by binding Factor H (FH), a complement regulator that inactivates C3b, preventing the formation of the C3-convertase and complement amplification on the bacterial surface. Factor H-related proteins (FHRs) are a group of plasma proteins evolutionarily related to FH that have been postulated to interfere in this bacterial mechanism of resisting complement. Here, we show that FHR-1 binds to P. aeruginosa via the outer membrane protein OprG in a lipopolysaccharide (LPS) O antigen-dependent manner. Binding assays with purified components or with FHR-1-deficient serum supplemented with FHR-1 show that FHR-1 competes with FH for binding to P. aeruginosa. Blockage of FH binding to C3b deposited on the bacteria reduces FH-mediated cofactor activity of C3b degradation, increasing the opsonization of the bacteria and the formation of the potent chemoattractant C5a. Overall, our findings indicate that FHR-1 is a host factor that promotes complement activation, facilitating clearance of P. aeruginosa by opsonophagocytosis.

Strongylocentrotus nudus Eggs Polysaccharide Enhances Macrophage Phagocytosis Against E.coli Infection by TLR4/STAT3 Axis.

Antibiotics resistance is one of the most significant public health threats globally. Strategies that strengthen host defenses to control pathogen infection has become a hot research field. Macrophages are part of early host defense mechanisms, and are activated via host pattern recognition receptors (PRRs), such as Toll-like receptor 4 (TLR4), which then facilitates phagocytosis and elimination of invading pathogens. However, few activators of PRRs have been approved for clinical use because of their toxic effects. This study aimed to investigate whether Strongylocentrotus nudus eggs polysaccharide (SEP), a non-toxic extract from seafood, contributes to host defense against bacterial infection. Results showed that SEP promoted bacterial clearance by enhancing phagocytosis by macrophages during E. coli infection in vitro, but was inhibited by TLR4 specific inhibitor TAK-242, STAT3 inhibitor Stattic or blockade of CD64. In addition, SEP protected mice from E. coli induced mortality, reduced pulmonary inflammation and inhibited dissemination of bacteria to organs, while TAK-242 retarded the protection of SEP. Overall, SEP strengthened innate host defense and improved the outcome in bacterial infection, suggesting that SEP could be used as a potential immunomodulator in host-directed therapies.

Attachment, penetration and early host defense mechanisms during the infection of filamentous brown algae by Eurychasma dicksonii.

Eurychasma dicksonii is one of the most common and widespread marine pathogens and attacks a broad spectrum of more than 45 brown algal species. The present study focuses on the mechanism used by the pathogen to attach on the host cell wall and force its way into algal cells. Ultrastructural examination revealed a needle-like structure which develops within the attached spore and extends along its main axis. Particular cell wall modifications are present at the basal part of the spore (adhesorium pad) and guide the needle-like tool to penetrate perpendicularly the host cell wall. The unique injection mechanism is shared with Haptoglossa species which suggests that this is an important characteristic of early diverging oomycetes. Furthermore, the encystment and adhesion mechanism of E. dicksonii shows significant similarities with other oomycetes, some of which are plant pathogens. Staining and immunolabelling techniques showed the deposition of β-1,3-glucans on the host cell wall at the pathogen penetration site, a strategy similar to physical responses previously described only in infected plant cells. It is assumed that the host defense in terms of callose-like deposition is an ancient response to infection.

Subproteome analysis of the neutrophil cytoskeleton

Neutrophils play a key role in the early host-defense mechanisms due to their capacity to migrate into inflamed tissues and phagocytose microorganisms. The cytoskeleton has an essential role in these neutrophil functions, however, its composition is still poorly understood. We separately analyzed different cytoskeletal compartments: cytosolic skeleton, phagosome membrane skeleton, and plasma membrane skeleton. Using a proteomic approach, 138 nonredundant proteins were identified. Proteins not previously known to associate with the skeleton were: n-acetylglucosamine kinase, phosphoglycerate mutase 1, prohibitin, ficolin-1, phosphogluconate dehydrogenase, glucosidase, transketolase, major vault protein, valosin-containing protein, aldehyde dehydrogenase, and lung cancer-related protein-8 (LCRP8). The majority of these proteins can be classified as energy metabolism enzymes. Such a finding was interesting because neutrophil energy metabolism is unusual, mainly relying on glycolysis. The enrichment of phosphoglycerate mutase in cytosolic skeleton was additionally indicated by the use of Western blotting. This is the broadest subcellular investigation to date of the neutrophil cytoskeletal proteome and the first proteomic analysis in any cell type of the phagosome skeleton. The association of metabolic enzymes with cytoskeleton is suggestive of the importance of their localized enrichment and macromolecular organization in neutrophils.

Early cellular responses in the peritoneal cavity of mice to antitumor immunomodulators.

The early cellular responses to antitumor immunomodulators and conventional inducers, especially the polymorphonuclear leukocyte (PMN) responses, were examined in the peritoneal cavity of mice to investigate their effect on primary defense mechanisms. Immunomodulators were classified into 5 groups in terms of PMN response on the basis of its duration (declining or persistent) and extent (high or low induction): 1) TAK (beta-1,3-glucan)-type (high, persistent), 2) lentinan-type (high, declining), 3) yeast mannan-type (low, declining), 4) LPS (lipopolysaccharide)-type (low, persistent), 5) others (no effect). Since the general PMN response is of the declining type, the persistence of PMN with TAK- and LPS-type immunomodulators is a characteristic of the PMN-inducing activity. With respect to the extent, TAK- and lentinan-type immunomodulators induced larger numbers of PMN and macrophages than conventional inducers. These results suggest that some types of immunomodulators have effects on the early host-defense mechanism. From the viewpoint of the general self-defense mechanism we also compared these PMN responses with those to bacteria and to tumor inoculation, and the properties of substances inducing high PMN response, i.e., those with the quality of "foreignness," are discussed.

Activation of phosphatidylinositol 3-kinase and c-Jun-N-terminal kinase cascades enhances NF-kappaB-dependent gene transcription in BCG-stimulated macrophages through promotion of p65/p300 binding.

The proinflammatory response of infected macrophages is an important early host defense mechanism against mycobacterial infection. Mycobacteria have been demonstrated to induce proinflammatory gene transcription through the Toll-like receptors, (TLR)2 and TLR 4, which initiate signaling cascades leading to nuclear factor (NF)-kappaB activation. The main transduction pathway responsible for NF-kappaB activation has been established and involves the MyD88, interleukin-1 receptor-associated kinase, tumor necrosis factor receptor-associated factor-6, NF-kappaB-inducing kinase, and inhibitor of kappaB kinase complex. The role of other kinase cascades triggered by mycobacteria in the NF-kappaB activation is less clear. We herein examine the role of the mitogen-activated protein kinases (MAPKs) and phosphatidylinositol 3-kinase (PI-3K) cascades in the expression of the bacillus Calmette-Guerin (BCG) mycobacteria-induced NF-kappaB-dependent genes, macrophage-inflammatory protein-2 (MIP-2) and inducible nitric oxide (NO) synthase. Specific pharmacological inhibition of the PI-3K, c-jun-N-terminal kinase (JNK), and to a smaller extent, p38 MAPK but not extracellular-regulated kinase (ERK), suppressed NF-kappaB-dependent reporter gene transcription and MIP-2 and NO secretion in BCG-induced RAW264.7 macrophages. A similar effect was obtained following molecular inhibition of JNK via JNK-interacting protein-1 overexpression. In addition, a kinase-dead mutant of MEK kinase-1, the up-stream regulator of JNK, also proved to be a potent inhibitor of NF-kappaB-reporter activity. The effect of inhibitors was mediated by the down-regulation of NF-kappaB transcription activity and without effecting its nuclear translocation. These data suggest an indirect mechanism of the NF-kappaB regulation by these kinases, probably through p65 phosphorylation and improved binding to the p300 transcription coactivator. The data obtained demonstrate that PI-3K, JNK, and p38 MAPK activation by mycobacteria enhance NF-kappaB-driven gene expression contributing to the proinflammatory macrophage response.

Chemokine-mediated recruitment of NK cells is a critical host defense mechanism in invasive aspergillosis.

Invasive aspergillosis is a severe pneumonia that is usually fatal despite currently available therapy. The disease disproportionately afflicts immunocompromised patients, indicating the critical importance of the immune status of the host in this infection, but the defense mechanisms against this pathogen remain incompletely understood. In the current study, we hypothesized that the chemokine ligand monocyte chemotactic protein-1, also designated CC chemokine ligand-2 (MCP-1/CCL2) is necessary for effective host defense against invasive aspergillosis in immunocompromised hosts. We found a rapid and marked induction of MCP-1/CCL2 in the lungs of neutropenic mice with invasive aspergillosis. Neutralizing MCP-1/CCL2 resulted in twofold greater mortality and greater than threefold increase in pathogen burden in the lungs. Neutralization of MCP-1/CCL2 also resulted in reduced recruitment of NK cells to the lungs at early time points, but did not affect the number of other leukocyte effector cells in the lungs. Ab-mediated depletion of NK cells similarly resulted in impaired defenses against the infection, resulting in a greater than twofold increase in mortality and impaired clearance of the pathogen from the lungs. These data establish MCP-1/CCL2-mediated recruitment of NK cells to the lungs as a critical early host defense mechanism in invasive aspergillosis and demonstrate NK cells to be an important and previously unrecognized effector cell in this infection.

Molecular analysis of the contribution of the capsular polysaccharide and the lipopolysaccharide O side chain to the virulence of Klebsiella pneumoniae in a murine model of pneumonia.

Klebsiella pneumoniae is a common cause of gram-negative bacterial nosocomial pneumonia. Two surface polysaccharides, lipopolysaccharide (LPS) O side chain and capsular polysaccharide (CPS), are critical for the microorganism in causing sepsis, but little is known about their role in pneumonia. To investigate their contribution in the pathogenesis of K. pneumoniae pneumonia, we characterized the host response to bacterial challenge with a highly virulent clinical isolate or with isogenic insertion-duplication mutants deficient in CPS or LPS O side chain in a murine model of pneumonia. Animals challenged intratracheally with the wild-type or LPS O side chain-deficient strain developed pneumonia and became bacteremic before death. Extensive lung lesions as well as pleuritis, vasculitis, and edema were observed by histopathological examination, and polymorphonuclear infiltration was also demonstrated. In contrast, none of the animals challenged with the unencapsulated strain developed pneumonia or bacteremia. Examination of tissue from this group did not identify lung lesions, and none of the infected animals died. Analysis of the early host defense mechanisms that contributed to the clearance of the unencapsulated mutant showed that the levels of C3 deposited on the unencapsulated mutant surface were threefold higher than those for the wild-type and LPS O side chain-deficient strains. Furthermore, phagocytosis of the unencapsulated mutant by human alveolar macrophages (AM) was more efficient than that of the wild-type and LPS O side chain-deficient strains. We conclude that CPS, but not LPS O side chain, is essential for Klebsiella pneumonia because it modulates the deposition of C3 and protects the microorganisms against human AM phagocytosis.

Kinetics of the early recruitment of leukocyte subsets at the sites of tumor cells in the lungs: natural killer (NK) cells rapidly attract monocytes but not lymphocytes in the surveillance of micrometastasis.

Early host defense mechanisms play a critical role for the outcome of metastatic disease but most of the initial steps of such responses against tumor cells are still unknown. Here, the specificity and kinetics of leukocyte subsets in response to intravenous inoculation of vital dye labeled Fischer 344 rat syngeneic MADB106 tumor cells were monitored in lungs in situ by immunohistochemistry and image analysis over a time-period of 6 hr. In comparison with sham injections, tumor cell inoculation induces a dynamic sequence of rapidly increasing granulocyte (+40% at 5 min), NK and T cell (+60% at 15 min) as well as monocyte (+100% at 30 min) numbers in lung tissue. Already within the first minutes frequent colocalizations of granulocytes and NK cells with tumor targets were found in situ. Within the first hour NK cells selectively kill tumor targets, because depletion of NK cells in vivo drastically increases both the number of MADB106 cells retained in lungs and the emerging numbers of lung tumor colonies. In addition, the tumor-cell-induced increase of monocytes strictly depends on the presence of NK cells because NK-depletion completely abrogates the time specific response of monocytes. Under NK depleted conditions the tumor-induced recruitment of CD4(+) T cells is more pronounced suggesting a compensatory mechanism. In contrast, B cell numbers progressively decrease within hours after cell inoculation. These findings demonstrate that NK and T cells mediate the initial steps in the surveillance of lung metastasis. NK cells rapidly kill tumor cells and subsequently recruit monocytes in vivo.

Immunologic aspects of eosinophilia.

There is compelling investigative data to indicate that the eosinophil may be more than just coincidentally involved in immune system function. In concert with IgE and the mast cell it is likely that eosinophil actions evolved as an early host defense mechanism against helminth parasites. Thus, a reasonable basis exists for the concept that in the evolutionary scheme there emerged genetically endowed humans with the potential for mounting immediate hypersensitivity responses to multicellular and complex antigenic particulates and animal and plant products, e.g., pollens and danders; hence the atopic state, allergic reactivity and the eosinophil. In many instances there is reason to interpret and ascribe a benign nature to eosinophil accumulations. In other circumstances, associations with hypersensitivity inflammation, for better or for worse, are viewed with special interest by eosinophil watchers as the unraveling of allergic phenomena continues.

HLA-restricted cytotoxic T lymphocytes are an early immune response and important defense mechanism in cytomegalovirus infections.

Eighty-eight bone marrow transplant recipients were studied for development of cytomegalovirus (CMV) infections and associated cytotoxic lymphocyte responses. Sixty-one patients developed CMV infection that was diagnosed by virus isolation (18 patients), fourfold rises in serum antibodies (13), or both (30). Interstitial pneumonitis developed in 27 patients. Among patients tested during infection, HLA-restricted, CMV-specific cytotoxic T cell responses occurred in 25 patients, and non-restricted, non-T cell cytotoxic responses, or responses of undetermined cell type, occurred in 10 patients. Levels of CMV-specific cytotoxicity in infected and uninfected patients were compared in two ways. Eighty-nine percent of patients (32 of 36) with cytotoxicity greater than or equal to 15% lysis (50:1 effector-to-target-cell ratio) and 100% of patients (21 of 21) with sequential increases in cytotoxicity of greater than or equal to 15% lysis were infected. The median time of occurrence of CMV-specific cytotoxic responses preceded the median onset of viral shedding by 1.4 weeks, or rises in titers of serum antibody by 1.7 weeks, and of interstitial pneumonitis by 2.5 weeks. Immune competence, as demonstrated by survival from infection, was always associated with a CMV-specific cytotoxic response. The HLA-restricted cytotoxic T cell response appears to be important as an early host-defense mechanism and an early diagnostic evidence of infection.

Macrophages in resistance to rickettsial infections: early host defense mechanisms in experimental scrub typhus.

Several early nonspecific host defense mechanisms were examined in resistant (BALB/c) and susceptible (C3H/He) mice after intraperitoneal inoculation with Rickettsia tsutsugamushi strain Gilliam. Inflammatory exudates were formed in both mouse strains in response to rickettsial inoculation, but the inflammatory response of C3H animals was delayed several days, and influx of peroxidase-positive macrophages occurred late in infection. Peritoneal cells of C3H mice became progressively infected, with 40% of both macrophages and lymphocytes containing intracellular rickettsiae by day 10. The early flammatory response of BALB/c mice was unexpectedly associated with a low percentage of infected peritoneal cells (1 to 2%). In vitro, no difference was detected in ability of resident macrophages of either strain to support the growth of R. tsutsugamushi or to become activated by treatment with lymphokines for rickettsiacidal activity. In vivo, however, macrophages from C3H mice inoculated with Gilliam were not activated on days 6 and 7 after infection, whereas BALB/c macrophages were continuously activated beginning on day 4. The lack of in vivo C3H macrophage activation was not secondary to deficient lymphokine production by infected lymphocytes, as levels of lymphokines produced by peritoneal lymphocytes of both strains were similar and peaked on day 7 after infection. Susceptibility to infection appears to be related to defective regulation of macrophage responses rather than to defects in macrophage function.