Effects of dimethyldioctadecylammonium bromide on phagocytosis and digestion of Listeria monocytogenes by mouse peritoneal macrophages.

Listeria monocytogenes was labelled with [3H]-thymidine and phagocytosis in vivo, measured after the intraperitoneal injection of killed or viable listeria. The loss of intracellular radioactivity after incubation in vitro was used as a measure for digestion. Killing was assessed by counting the numbers of viable listeria before and after in vitro incubation. Peritoneal macrophages from mice immunized with viable listeria showed better phagocytosis and digestion of both killed and viable listeria than macrophages from normal mice. Viable listeria were digested to a lesser degree than killed ones by both normal and immune macrophages. The simultaneous injection of the surfactant dimethyldioctadecylammonium bromide (DDA) with killed or viable listeria greatly depressed the digestion of listeria. Phagocytosis of viable listeria was not affected whereas that of killed listeria was slightly enhanced. The injection of 10(8) killed listeria mixed with DDA greatly impaired the digestion of viable listeria 7 days but not 1 day after injection. The impairment of digestion was accompanied by an increase in the number of intracellular viable listeria.     

Effects of BCG infection on the susceptibility of mouse macrophages to endotoxin.

Mice infected intravenously with Mycobacterium bovis (BCG) are 100 to 1,000 times more sensitive to the lethal effects of bacterial lipopolysaccharides (LPS). Since BCG infection results in macrophage activation and LPS may cause pathophysiological effects through interaction with this cell type, it was of interest to determine whether macrophages from BCG-infected animals were more susceptible to the toxic effects of LPS in vitro. When LPS-susceptible, C57BL/6 mice were infected with BCG, a significant reduction in the 50% lethal dose of LPS was first observed after 7 days and persisted for several weeks. Macrophages from these animals had greatly increased susceptibility to LPS in vitro, which correlated with the development of acquired cellular resistance as determined by their ability to inhibit the growth of Listeria monocytogenes. In contrast, BCG infection of C3H/HeJ mice, a strain resistant to LPS, did not alter the 50% lethal dose of LPS for these animals or increase the sensitivity of their peritoneal macrophages to LPS in vitro. These results indicate that susceptibility of BCG-infected mice to the lethal effects of LPS parallels the susceptibility of their macrophages in vitro; release of vasoactive substances from LPS-susceptible activated macrophages in vivo may be, in part, responsible for lethality.     

Growth of Mycobacterium avium in activated macrophages harvested from inbred mice with differing innate susceptibilities to mycobacterial infection.

The growth of Mycobacterium avium in macrophages obtained from Mycobacterium bovis BCG-infected mice was compared with that in macrophages from uninfected mice. BCG vaccination resulted in substantial macrophage activation, measured as increased acid phosphatase and superoxide anion production, as well as enhanced leishmanicidal activity. However, the activated macrophages were only able to reduce the rate of intracellular growth by Listeria monocytogenes and M. avium in vivo and did not express detectable levels of mycobactericidal activity in vitro. Exposure of the macrophage monolayers to concanavalin A-stimulated spleen cell supernatant fluid and lipopolysaccharide did not further enhance the ability of the BCG-activated macrophages to control the intracellular replication of the M. avium. Macrophages from BCG-infected C57BL/6 (BCGs) mice were quantitatively better able to control the intracellular replication of the M. avium challenge than were similar phagocytes obtained from BCGr (A/J) mice. These findings have important implications with respect to the expression of acquired resistance to these atypical mycobacterial infections.     

Host defenses in murine malaria: nonspecific resistance to Plasmodium berghei generated in response to Mycobacterium bovis infection or Corynebacterium parvum stimulation.

Infection with Mycobacterium bovis (BCG) or injection of killed Corynebacterium parvum protected some strain B6D2 F1 (C57BL/6xDBA/2) mice but did not protect strain ICR or A mice from lethal challenge with Plasmodium berghei strain NYU-2. B6D2 mice were not protected against challenges delivered immediately after intravenous injection of these materials, but rather protection developed by day 7 and persisted through at least day 84. Infections in protected mice progressed to about 10% parasitemia in parallel with infections initiated with the same inoculum in untreated controls. However, infections in most of the protected mice were cleared subsequently, whereas infections in untreated controls were uniformly fatal. A small number of treated mice developed protracted high-level erythrocytic infections, which led to markedly delayed death. BCG-infected mice which survived P. berghei infections had a factor in their sera which protected passively immunized recipients from P. berghei. BCG-infected mice passively immunized with protective serum controlled P. berghei infections better than normal mice given the same amount of the same serum and challenged with the same P. berghei inoculum. The capacity of BCG-infected B6D2 mice to resist P. berghei infection was not directly related to the pattern of growth of BCG, to the degree of splenomegaly, or to the level of activation of macrophages (measured as microbicidal capacity) caused by BCG infection. Therefore, I concluded that (i) BCG infection or injection of killed C. parvum altered the immunological potential of B6D2 mice in such a way as to allow the production of measurable levels of a protective humoral factor in response to infection with P. berghei; (ii) BCG infection caused the generation of a capacity which, when expressed in the presence of immune serum, provided an anti-P. berghei capacity which was superior to that provided by BCG infection alone or immune serum in the absence of BCG infection; and (iii) not all strains of mice could be protected from P. berghei by BCG or C. parvum injection.     

Effect of Dihydrostreptomycin on Phagocytosis of Mouse-Peritoneal Macrophages In Vitro

The effect of the antibiotic dihydrostreptomycin on the phagocytic and bactericidal ability of peritoneal macrophages obtained from mice has been investigated. In subliminal concentrations which did not influence the bacterial growth, the drug caused macrophages to ingest and kill bacteria (Escherichia coli) at a higher rate than did macrophages without antibiotic. The differences for phagocytosis and intracellular killing of E. coli with and without a subliminal amount of dihydrostreptomycin were statistically significant. Macrophages pretreated with the antibiotic did not demonstrate any enhancement of phagocytosis.     

Malaria-induced lymphokines: stimulation of macrophages for enhanced phagocytosis

Culture supernatants from antigen-pulsed spleen cells of mice infected previously with either BCG or Plasmodium chabaudi were used to study macrophage activation as judged by phagocytosis of immunoglobulin G-sensitized erythrocytes and Plasmodium berghei- and P. chabaudi-infected erythrocytes. Resident peritoneal macrophages were incubated in vitro with spleen cell factor and then assayed for ingestion of immunoglobulin G-sensitized or parasitized erythrocytes. Macrophages activated with BCG-induced lymphokine bound and ingested two- to threefold more P. berghei parasitized erythrocytes than macrophages incubated with control spleen cell factor. Similarly, Plasmodium-stimulated spleen cells from mice infected with malaria produced a lymphokine(s) capable of activating macrophages for enhanced Fc receptor-mediated phagocytosis. The stimulation of phagocytosis by the lymphokine is nonspecific in nature, since phagocytosis of parasitized erythrocytes from one species of murine malaria is enhanced by the lymphokine prepared from a heterologous species. Nylon wool-nonadherent, malaria-sensitized spleen cells elaborated a lymphokine which stimulates macrophages for enhanced phagocytosis, whereas anti-0-treated spleen cells failed to produce the phagocytosis-promoting lymphokine. Consequently, this lymphokine appears to be elaborated by sensitized T lymphocytes. Interestingly, enhanced phagocytosis of opsonized trophozoites and schizonts, but not ring stage parasites of P. chabaudi, was displayed by macrophages activated with the lymphokine(s) prepared from P. chabaudi-recovered mice. Preincubation of the malaria parasitized erythrocytes with hyperimmune serum raised against the parasites greatly facilitated both binding and ingestion by the stimulated macrophages.     

Interaction of Alveolar Macrophages with Nocardia asteroides: Immunological Enhancement of Phagocytosis, Phagosome-Lysosome Fusion, and Microbicidal Activity

Normal and specifically activated rabbit alveolar macrophages were infected in vitro with Nocardia asteroides GUH-2. In the presence of serum from normal rabbits, no significant differences were noted between normal and activated alveolar macrophages with respect to phagocytosis, incidence of phagosomelysosome fusion, or nocardicidal activity. However, all of these macrophage functions were enhanced by various immunological components. Serum from immunized rabbits enhanced phagocytosis of nocardial cells by activated macrophages, and there was an additional increase in phagocytosis observed when alveolar lining material was present. Complement had no effect on the ability of the macrophages to phagocytize nocardial cells. The greatest percentage of organisms phagocytized was observed when specifically primed lymph node cells, alveolar lining material, and serum from immunized rabbits were present in the incubation medium. N. asteroides GUH-2 inhibited phagosome-lysosome fusion in normal macrophages in the presence of serum from normal rabbits. However, addition of serum from immunized rabbits or the addition of specifically primed lymphocytes increased the amount of phagosome-lysosome fusion, whereas complement had no effect on this fusion process. Nocardial viability was not reduced when either normal or activated macrophages were infected with bacteria in the presence of normal serum, immune serum, or alveolar lining material. However, specifically activated macrophages incubated with primed lymph node cells obtained from immunized rabbits were able to both decrease the number of viable organisms recovered and to increase the incidence and extent of bacterial cell damage. The greatest number of organisms were killed by specifically activated macrophages when the bacterial cells were incubated with primed lymph node cells suspended in immune serum and alveolar lining material. These results indicate that activated macrophages alone are not sufficient to kill ingested N. asteroides GUH-2 and that specifically primed lymphocytes are important in host resistance to nocardial infections.     

BCG-induced enhancement of endotoxin sensitivity in C3H/HeJ mice. II. T cell modulation of macrophage sensitivity to LPS in vitro

BCG infection induces a marked increase in LPS sensitivity in vivo and will render genetically defective, LPS hyporesponsive, C3H/HeJ mice almost as sensitive to LPS as normal mice. In this study, we have examined the endotoxin sensitivity of lymphocytes and macrophages from BCG infected mice in order to determine the cellular basis of this effect. We have found that the alteration in endotoxin sensitivity is mediated by a primary effect of BCG infection on T lymphocytes rather than on macrophages. Macrophages from «LPS sensitive», BCG-infected C3H/HeJ mice remain unresponsive to LPS when tested in vitro. However, when peritoneal T lymphocytes from these LPS »corrected» mice were cocultured with LPS unresponsive C3H/HeJ macrophages, a conversion to the LPS-responsive state was observed as manifested by the ability of the macrophages to produce LAF (IL 1) upon LPS stimulation. T cells from normally LPS-responsive or BCG-infected C3H/HeN mice, but not from control C3H/HeJ mice, were also able to render C3H/HeJ macrophages sensitive to LPS. This activity was not affected by treatment of the column-purified T cells with anti-macrophage serum plus complement, indicating that the response was not due to residual LPS-responsive macrophages contaminating the T cell preparations. The ability of the T cell suspension to render C3H/HeJ macrophages capable of producing LAF (IL 1) in response to LPS was abrogated by treatment of the T cell preparations with anti-Thy 1.2 plus complement. These findings establish the importance of T lymphocytes in regulating the LPS sensitivity of macrophages in BCG infected C3H/HeJ mice and support the concept that macrophage LPS responsiveness is dependent upon a certain state of macrophage activation which is regulated by lymphocytes.     

Neutralization of gamma interferon and tumor necrosis factor alpha blocks in vivo synthesis of nitrogen oxides from L-arginine and protection against Francisella tularensis infection in Mycobacterium bovis BCG-treated mice.

Peritoneal cells from Mycobacterium bovis BCG-infected C3H/HeN mice produced nitrite (NO2-, an oxidative end product of nitric oxide [NO] synthesis) and inhibited the growth of Francisella tularensis, a facultative intracellular bacterium. Both NO2- production and inhibition of bacterial growth were suppressed by NG-monomethyl-L-arginine, a substrate inhibitor of nitrogen oxidation of L-arginine, and monoclonal antibodies (MAbs) to gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha). Intraperitoneal injection of mice with BCG increased urinary nitrate (NO3-) excretion coincident with development of activated macrophages capable of secreting nitrogen oxides and inhibiting F. tularensis growth in vitro. Eight days after BCG inoculation, mice survived a normally lethal intraperitoneal challenge with F. tularensis. Treatment of these BCG-infected mice with MAbs to IFN-gamma or TNF-alpha at the time of BCG inoculation reduced urinary NO3- levels to those found in normal uninfected mice for up to 14 days. The same anticytokine antibody treatment abolished BCG-mediated protection against F. tularensis: mice died within 4 to 6 days. Intraperitoneal administration of anti-IFN-gamma or anti-TNF-alpha antibody 8 days after BCG infection also reduced urinary NO3- and abolished protection against F. tularensis. Isotype control (immunoglobulin G) or anti-interleukin 4 MAbs had little effect on these parameters at any time of treatment. IFN-gamma and TNF-alpha were clearly involved in the regulation of macrophage activation by BCG in vivo. Protection against F. tularensis challenge by BCG depended upon the physiological generation of reactive nitrogen oxides induced by these cytokines.     

Serum amyloid P-component-induced enhancement of macrophage listericidal activity.

Purified serum amyloid P component (SAP), the major acute-phase reactant of mice, augmented the in vitro listericidal activity of inflammatory (elicited) macrophages, bone marrow-derived monocytes, and macrophages from a subcutaneous site of inflammation. Monocytes and macrophages from C57BL/B6 mice, which are relatively resistant to Listeria monocytogenes, exhibited a significantly greater enhanced killing capacity for listeria than macrophages from listeria-susceptible A/J mice. SAP did not alter the extent of phagocytosis by macrophages of opsonized L. monocytogenes, nor was SAP opsonic for listeria. Mannose-derived simple sugars inhibited the binding of SAP to macrophages and consequently prevented the enhanced SAP-dependent listericidal activity. Macrophages from lipopolysaccharide-hyporesponsive mice also had increased microbicidal activity following incubation with SAP. SAP activated macrophages independently of lymphokine. Therefore, SAP may serve as a mediator of the heightened nonspecific host defense response that is associated with the acute phase of the systemic inflammatory response.     

Rapid and complete fusion of macrophage lysosomes with phagosomes containing Salmonella typhimurium.

The virulence of Salmonella typhimurium for mice results, in part, from its ability to survive after phagocytosis by macrophages. Although it is generally agreed that intracellular bacteria persist in membrane-bound phagosomes, there remains some question as to whether these phagosomes fuse with macrophage lysosomes. This report describes the maturation of phagosomes containing S. typhimurium inside mouse bone marrow-derived macrophages. Macrophages were infected briefly and incubated for various intervals; then they were examined by fluorescence microscopy for colocalization of bacteria with lysosomal markers. These markers included LAMP-1, cathepsin L, and fluorescent proteins or dextrans preloaded into lysosomes by endocytosis. By all measures, phagosomes containing S. typhimurium merged completely with the lysosomal compartment within 20 min of phagocytosis. The rate of phagosome-lysosome fusion was similar to the rate for phagocytosed latex beads. Phagolysosomes remained accessible to fluid-phase probes and contained lysosomal markers for many hours. Moreover, a large percentage of the wild-type bacteria that were viable 20 min after infection survived longer incubations inside macrophages, indicating that the survivors were not a minor subpopulation that avoided phagosome-lysosome fusion. Therefore, we conclude that S. typhimurium survives within the lysosomal compartments of macrophages.     

Enhanced elimination of Listeria monocytogenes at the site of delayed footpad reaction.

The protective activity against a challenge infection with Listeria monocytogenes was investigated at the site of a delayed footpad reaction in mice immunized with viable or killed listeria. Delayed footpad reactivity was induced only in mice immunized with viable bacteria. Rapid and marked elimination of challenge bacteria was observed only at the site of reaction in mice immunized with viable bacteria but not in mice immunized with killed bacteria. Macrophage migration inhibitory activity was observed equally in both groups of mice. These results suggest that the delayed footpad reaction contributes directly to the elimination of bacteria irrespective of macrophage migration inhibitory activity.     

The role of macrophages in the protection of mice against leptospirosis: in vitro and in vivo studies

Balb/c mice are naturally resistant to infection with Leptospira interrogans serovar copenhageni, but leptospires were not phagocytosed by mouse peritoneal macrophages in vitro without added specific antibody. Similar results were obtained irrespective of whether leptospires were viable or killed, virulent or avirulent, or whether macrophages were obtained from normal mice, immunized mice or mice previously infected with BCG. Suppression or stimulation of macrophage function in vivo did not affect the outcome of infection of immunosuppressed mice with leptospires; specific antibody was essential for protection from infection.     

Superoxide production in pulmonary alveolar macrophages and killing of BCG by the superoxide-generating system with or without catalase.

The superoxide production of BCG-infected and noninfected alveolar macrophages was measured by superoxide dismutase-inhibitable nitro blue tetrazolium reduction. The cells were incubated with or without cell-free bronchial lavage fluid (pulmonary washings). When control alveolar macrophages were infected by BCG, superoxide production was decreased markedly, probably due to bacterial cytotoxic factors. In contrast, the production of superoxide in alveolar macrophages exposed to pulmonary washings was increased and not appreciably influenced by BCG infection. Superoxide production by alveolar macrophages was dependent on time and on the protein concentration in the pulmonary washings. In controls, it was inversely proportional to the infecting dose of BCG. We observed previously that alveolar macrophages activated by pulmonary washings inhibited intracellular growth of BCG. We now present evidence that enhanced production of superoxide contributes to such inhibition, especially in the presence of catalase at acid pH. These findings are pertinent to the defense of inflamed lungs, where serum and serum immunoglobulin G transuded from blood into alveolar spaces probably induce such activation on alveolar macrophages.     

Natural killer cell activity and macrophage-dependent inhibition of growth or killing of Mycobacterium avium complex in a mouse model

Natural killer (NK) cells from spleens of normal and Mycobacterium avium complex (MAC)-infected C57 black mice (C57 BL/6 bg/+) were examined for their capacity to activate splenic and peritoneal macrophages from beige mice to inhibit or kill intracellular MAC. Peritoneal and splenic macrophages from beige mice were exposed in vitro to NK cells obtained from MAC-infected and uninfected black mice. NK cells from uninfected black mice were also treated in vitro with recombinant interleukin-2 (IL-2) for 48 h before incubation with macrophages. While control macrophages supported intracellular growth of MAC, macrophages exposed to unactivated NK cells inhibited growth of the intracellular bacteria, as determined 4 days after infection. IL-2 stimulated NK cells, and NK cells obtained from MAC-infected animals were able to activate murine macrophages in vitro to inhibit growth or kill 40.0 +/- 5% and 61.3 +/- 6% of the intracellular bacteria, respectively. In other experiments, beige mice (C57 BL/6 bg/bg) were treated intraperitoneally with NK cells obtained from MAC-infected and uninfected C57 black mice. Peritoneal macrophages harvested from beige mice treated with NK cells activated in vitro with IL-2 killed 24.4 +/- 4% of intracellular bacteria by day 4 after infection. Macrophages obtained from animals treated with NK cells harvested from MAC-infected black mice killed 58.8 +/- 7% of intracellular bacteria by 4 days after infection, in contrast with intracellular growth observed in macrophages obtained from untreated animals and from animals treated with Hanks' solution or unactivated NK cells. These crossover studies suggest that NK cells may be important in host defense against MAC.     

TLR2 and TLR4 serve distinct roles in the host immune response against Mycobacterium bovis BCG

Toll-like receptor (TLR) proteins mediate cellular activation by microbes and microbial products. To delineate the role of TLR proteins in the development of host immune responses against mycobacteria, wild-type and TLR-deficient mice were infected with nonpathogenic Mycobacterium bovis bacillus Calmette-Guerin (BCG). Two weeks after intraperitoneal challenge with BCG, few bacilli were present in the lungs of wild-type and TLR4(-/-) mice, whereas bacterial loads were tenfold higher in the lungs of infected TLR2(-/-) mice. BCG challenge in vitro strongly induced proinflammatory cytokine secretion by macrophages from wild-type and TLR4(-/-) mice but not by TLR2(-/-) macrophages. In contrast, intracellular uptake, intracellular bacterial growth, and suppression of intracellular bacterial growth in vitro by interferon-gamma (IFN-gamma) were similar in macrophages from all three mouse strains, suggesting that BCG growth in the lungs of TLR2(-/-) mice was a consequence of defective adaptive immunity. Antigenic stimulation of splenocytes from infected wild-type and TLR4(-/-) mice induced T cell proliferation in vitro, whereas T cells from TLR2(-/-) mice failed to proliferate. Unexpectedly, activated CD4(+) T cells from both TLR-deficient mouse strains secreted little IFN-gamma in vitro compared with control T cells. A role for TLR4 in the control of bacterial growth and IFN-gamma production in vivo was observed only when mice were infected with higher numbers of BCG. Thus, TLR2 and TLR4 appear to regulate distinct aspects of the host immune response against BCG.     

Fate of Legionella pneumophila Philadelphia-1 strain in resident, elicited, activated, and immune peritoneal macrophages of guinea pigs.

Legionella pneumophila is known to grow intracellularly in resident peritoneal macrophages of guinea pigs. The present study was done to determine what kinds of macrophage stimulants are able to activate guinea pig macrophages to inhibit intracellular growth of the organism. Peritoneal macrophages were harvested from healthy guinea pigs, from guinea pigs injected intraperitoneally with proteose peptone (PP) or thioglycolate medium, from guinea pigs injected intraperitoneally with live Mycobacterium bovis BCG or killed Propionibacterium acnes (Corynebacterium parvum), and from guinea pigs surviving infection with live L. pneumophila. After in vitro phagocytosis, the L. pneumophila CFU in each well were counted on charcoal-yeast extract agar plates. In the macrophages elicited by PP or thioglycolate medium, the organism grew as well as it did in resident macrophages. In BCG-activated and immune macrophages, growth was inhibited almost completely. In P. acnes-activated macrophages, the initial growth of L. pneumophila was inhibited to some extent, but its growth reached the same level as in the resident and PP-induced macrophages after 3 or 4 days of culture. In the lethal challenge experiments in vivo, the superior protection provided by BCG over P. acnes was ascertained and the importance of macrophages in resistance to L. pneumophila was confirmed. Difference of activation by BCG and P. acnes in relation to the inhibition of intracellular growth of L. pneumophila in guinea pig macrophages is discussed.     

Infection with Mycobacterium bovis BCG diverts traffic of myelin oligodendroglial glycoprotein autoantigen-specific T cells away from the central nervous system and ameliorates experimental autoimmune encephalomyelitis

Infectious agents have been proposed to influence susceptibility to autoimmune diseases such as multiple sclerosis. We induced a Th1-mediated central nervous system (CNS) autoimmune disease, experimental autoimmune encephalomyelitis (EAE) in mice with an ongoing infection with Mycobacterium bovis strain bacillus Calmette-Guérin (BCG) to study this possibility. C57BL/6 mice infected with live BCG for 6 weeks were immunized with myelin oligodendroglial glycoprotein peptide (MOG(35-55)) to induce EAE. The clinical severity of EAE was reduced in BCG-infected mice in a BCG dose-dependent manner. Inflammatory-cell infiltration and demyelination of the spinal cord were significantly lessened in BCG-infected animals compared with uninfected EAE controls. ELISPOT and gamma interferon intracellular cytokine analysis of the frequency of antigen-specific CD4(+) T cells in the CNS and in BCG-induced granulomas and adoptive transfer of MOG(35-55)-specific green fluorescent protein-expressing cells into BCG-infected animals indicated that nervous tissue-specific (MOG(35-55)) CD4(+) T cells accumulate in the BCG-induced granuloma sites. These data suggest a novel mechanism for infection-mediated modulation of autoimmunity. We demonstrate that redirected trafficking of activated CNS antigen-specific CD4(+) T cells to local inflammatory sites induced by BCG infection modulates the initiation and progression of a Th1-mediated CNS autoimmune disease.     

Neutrophil secretion products regulate anti-bacterial activity in monocytes and macrophages

Macrophages represent a multi-functional cell type in innate immunity that contributes to bacterial clearance by recognition, phagocytosis and killing. In acute inflammation, infiltrating neutrophils release a wide array of preformed granule proteins which interfere functionally with their environment. Here, we present a novel role for neutrophil-derived granule proteins in the anti-microbial activity of macrophages. Neutrophil secretion obtained by antibody cross-linking of the integrin subunit CD18 (X-link secretion) or by treatment with N-Formyl-Met-Leu-Phe (fMLP secretion) induced a several-fold increase in bacterial phagocytosis by monocytes and macrophages. This response was associated with a rapid activation of the monocytes and macrophages as depicted by an increase in cytosolic free Ca(2+). Interestingly, fMLP secretion had a more pronounced effect on monocytes than the X-link secretion, while the opposite was observed for macrophages. In addition, polymorphonuclear cells (PMN) secretion caused a strong enhancement of intracellular reactive oxygen species (ROS) formation compared to incubation with bacteria. Thus, secretion of neutrophil granule proteins activates macrophages to increase the phagocytosis of bacteria and to enhance intracellular ROS formation, indicating pronounced intracellular bacterial killing. Both mechanisms attribute novel microbicidal properties to PMN granule proteins, suggesting their potential use in anti-microbial therapy.