Lowering of bactericidal activity of mouse peritoneal macrophages under combined use of staphylococcal enterotoxin type A and endotoxin

The present study investigated the effect of staphylococcal enterotoxin type A (SEA) and endotoxin Serratia marcescens (LPS) on the phagocytosis and killing of Staphylococcus aureus by mouse peritoneal macrophages. Two hours after enterotoxin intraperitoneal injection phagocytic and bactericidal activity were depressed. 24 hours later there was increased functional activity of macrophages by SEA and LPS, apart. But when two toxins were administered together (LPS four hours later enterotoxin) marked inhibition of bacterial killing was observed. When peritoneal macrophages were treated in vitro for 24 hours with the same toxins they were also markedly suppressed in bactericidal activity.     

Decreased outer membrane permeability protects mycobacteria from killing by ubiquitin-derived peptides

Ubiquitin-derived peptides are bactericidal in vitro and contribute to the mycobactericidal activity of the lysosome. To further define interactions of ubiquitin-derived peptides with mycobacteria, we screened for mutants with increased resistance to the bactericidal activity of the synthetic ubiquitin-derived peptide Ub2. The four Ub2-resistant Mycobacterium smegmatis mutants were also resistant to the bactericidal action of other antimicrobial peptides and macrophages. Two mutants were in the mspA gene encoding the main M. smegmatis porin. Using a translocation-deficient MspA point mutant, we showed that susceptibility of M. smegmatis to Ub2 was independent of MspA channel activity. Instead, the M. smegmatis Ub2-resistant mutants shared a common phenotype of decreased cell wall permeability compared with wild-type bacteria. Expression of mspA rendered Mycobacterium tuberculosis CDC1551 more susceptible both to ubiquitin-derived peptides in vitro and to lysosomal killing in macrophages. Finally, biochemical assays designed to assess membrane integrity indicated that Ub2 treatment impairs membrane function of M. smegmatis and M. tuberculosis cells . The M. smegmatis Ub2-resistant mutants were more resistant than wild-type M. smegmatis to this damage. We conclude that Ub2 targets mycobacterial membranes and that reduced membrane permeability provides mycobacteria intrinsic resistance against antimicrobial compounds including bactericidal ubiquitin-derived peptides.     

Effect of iron and phagocytosis on murine macrophage activation in vitro

Iron-exposed murine macrophages have a modified bactericidal activity as shown by previous observations. In order to assess the role of iron in macrophage activation, as measured by free radical production and by intracellular bacterial killing, murine peritoneal macrophages were cultivated in the presence of various sources of iron, human iron-saturated transferrin and ammonium ferric citrate, or iron chelators, Desferal, and human Apo-transferrin, and were infected with an enteropathogenic strain ofE. coli. The release of nitrite (NO₂ ^?), and the production of superoxide anion (O₂ ^?) and hydrogen peroxide (H₂O₂) by the phagocytes were measured and compared to the production by uninfected macrophages. The synergistic action with murine r.IFN-γ was also studied in the radical production reaction and for the bactericidal activity of macrophages. Our results show that in vitro phagocytosis ofE. coli induced elevated production of NO₂ ^? and H₂O₂ by macrophages, and that oxygen derivatives were released independently of the presence of added iron or chelator. Despite a phagocytosis-related enhancement of NO₂ ^? release, reactive nitrogen intermediates (RNI) are not directly involved in the bactericidal mechanism, as revealed by increased intracellular killing owing to RNI inhibitors. Moreover, bacterial killing may depend on oxygen derivatives, as suggested by the effect of the antioxidant sodium ascorbate leading to both a diminished H₂O₂ production and a decreased bactericidal activity of macrophages.     

Interferon enhances the bactericidal activity of peritoneal exudate macrophages against Salmonella typhimurium in mice

The treatment of macrophages of mouse peritoneal exudate has been found to enhance their bactericidal activity with respect to S. typhimurium. This activation depends on the dose of interferon and the cells/bacteria ratio. The action of interferon is species-specific.     

Determination of the bactericidal strength of alveolar macrophages by using the NBT test

The NBT test on alveolar macrophages can be used for the indirect evaluation of their bactericidal potency. This test has made it possible to show that the development of the tuberculous process in the lungs is characterized by the increasing intensity of the reduction of NBT by alveolar macrophages, which indirectly indicates their considerable bactericidal potency. But 3-4 weeks after infection macrophages, though capable of considerable bactericidal action, do not react to stimulation by Mycobacterium tuberculosis.     

Killing of Escherichia coli by mononuclear phagocytes and neutrophils stimulated in vitro with beta-1,3-D-polyglucose derivatives.

Human monocytes, human peritoneal macrophages, mouse peritoneal macrophages and human peripheral neutrophils pretreated with beta-1,3-D-polyglucose derivatives showed pronounced bactericidal capacity to Escherichia coli compared to control cells. The increased bactericidal capacity was detectable in mononuclear phagocytes over a wide range of concentrations of bacteria. Granulocytes, however, showed bactericidal capacity only at low concentrations of bacteria. The pretreated mononuclear phagocytes released significant amounts of IL-1 and PGE2. However, there was no significant release of tumor necrosis factor (TNF). By incubating unstimulated cells with purified IL-1 and TNF, the bactericidal activity of neutrophils and mononuclear phagocytes was enhanced. Our data indicate that the inability of neutrophils stimulated with beta-1,3-D-polyglucose derivatives to kill large numbers of bacteria could be overcome by a combined treatment with purified IL-1 or TNF in addition to beta-1,3-D-polyglucose derivatives. By incubating unstimulated cells with medium from beta-1,3-D-polyglucose-treated human peritoneal macrophages, the bactericidal activity of the cells was enhanced to the same extent as cells pretreated with purified TNF and IL-1. Cells incubated with IL-1-depleted medium from beta-1,3-D-polyglucose-treated human peritoneal macrophages, showed reduced bactericidal activity compared to cells incubated with undepleted medium. These studies demonstrate that beta-1,3-D-polyglucose-treated mononuclear phagocytes and neutrophils show enhanced bactericidal activity. The enhanced activity is partly caused by stimulation of the cells with IL-1 released from mononuclear phagocytes and partly by other unknown effects of beta-1,3-D-polyglucose derivatives on both mononuclear phagocytes and neutrophils.     

The heterogeneity of the alveolar macrophages of guinea pigs immunized subcutaneously with a vaccinal strain of Yersinia pestis]

The study of alveolar macrophages obtained from guinea pigs at different periods after the subcutaneous immunization with Y. pestis vaccine strain has revealed the heterogeneity of these macrophages in their bactericidal activity with respect to Y. pestis and in their resistance to the cytopathic action of this microorganism, as well as in the content of lysosomes and in the expression of receptors on the cell surface. The study has shown that subcutaneous immunization does not lead to the redistribution of subpopulations and to changes in their digestive activity.     

Antibody increases phagocytosis and killing of Lactococcus garvieae by rainbow trout (Oncorhynchus mykiss, L.) macrophages

The present study reports that specific antibody increased the bactericidal activity of rainbow trout head-kidney macrophages against virulent capsulated Lactococcus garvieae in the absence of complement. The observed increased bactericidal activity appeared to result from increased phagocytosis of capsulated L. garvieae in the presence of specific antibody and may in part explain the protective effect of antibody previously reported against this disease.     

Phagocytic activity of peripheral blood monocytes and peritoneal macrophages in the progeny of animals with experimental chronic liver disease

Peripheral blood monocytes and peritoneal macrophages (phagocytic index, phagocytosis intensity, metabolic level) in the offspring of mice with chronic experimental autoimmune liver affection have been studied for different parameters of their phagocytic properties. The obtained results testify to absorption and bactericidal activity disturbance of mononuclears studied in this group of animals.     

Enhancement of bactericidal activity of mouse peritoneal macrophages against Staphylococcus aureus by mouse interferon preparations

The effect of mouse interferon on the bactericidal activity of macrophages against pyogenic cocci was examined. Mouse peritoneal macrophages were cultivated with Staphylococcus aureus in vitro and viable Staphylococcus was recovered by treatment of the mixed macrophage-bacteria culture with sodium dodecyl sulphate (SDS) solution. Results showed that S. aureus was phagocytized and killed by the macrophages. Mouse L cell interferon enhanced the bactericidal activity of macrophages. A mouse brain interferon preparation also enhanced this activity. However, heat-inactivated L cell interferon and heterologous rabbit RK-13 cell interferon and human leukocyte interferon did not enhance it. This suggests that interferon enhances the bactericidal activity of macrophages against S. aureus.     

Changes in the bactericidal activity of macrophages from mice with various degrees of resistance to Salmonella typhimurium infection as affected by interferon

The bactericidal activity of mouse macrophages with different sensitivity to Salmonella infection has been studied. The sensitivity of BALB/c mice to S. typhimurium infection is associated with the low bactericidal activity of their macrophages. The introduction of interferon stimulates the bactericidal activity of macrophages sensitive to Salmonella infection of mice, which sharply enhances the resistance of the animals to this infection.     

Repeated social defeat increases the bactericidal activity of splenic macrophages through a Toll-like receptor-dependent pathway

Phagocytes of the innate immune system, such as monocytes/macrophages, represent a first line of defense against invading microorganisms. Psychological stress is often thought to suppress the functioning of these cells, in part due to the immunosuppressive activity of stress-induced glucocorticoid hormones. However, exposure to the stressor social disruption (SDR) has been shown to increase cytokine production by monocytes/macrophages and to reduce their sensitivity to corticosterone. Thus, it was hypothesized that splenic monocytes/macrophages from socially stressed mice would be primed to be more physiologically active than cells from nonstressed controls. Flow cytometry was used to demonstrate that exposure to SDR significantly increased the expression of Toll-like receptors (TLR) 2 and 4 on the surface of splenic macrophages. In a follow-up experiment, exposure to SDR also increased the ability of these macrophages to kill Escherichia coli ex vivo and in vivo. However, SDR failed to increase the bactericidal activity of splenic macrophages from C3H/HeJ mice, which lack functional TLR4. In mice with functional TLR4, the stress-induced increase in bactericidal activity was associated with a significant increase in macrophage gene expression for inducible nitric oxide synthase and subunits of the NADPH oxidase complex, which are responsible for generating reactive nitrogen and oxygen intermediates, respectively. This stress-induced increase in gene expression was not evident in the TLR4-deficient mice. These data indicate that SDR increases TLR expression, which in turn enhances the bactericidal activity of splenic macrophages, in part by increasing pathways responsible for reactive oxygen and nitrogen intermediate production.     

Effect of pertussis vaccine on the functional activity of the peritoneal and splenic macrophages in 2 mouse strains genetically differing by the intensity of their immune response

The immunostimulating effect of corpuscular pertussis vaccine on the antigen-presenting and bactericidal functions of peritoneal and splenic macrophages in CBA and C57BL/6 mice, differing in the intensity of immune response to sheep red blood cells and Salmonella typhimurium, has been studied. The study has revealed that the injection of pertussis vaccine alters the functional activity of the cells under study, the effect depending on the immunizing dose, the strain of mice and the time elapsed from the moment of immunization. Pertussis vaccine enhances the low capacity of macrophages for antigen presentation in C57BL/6 mice with low responsiveness and alters the resistance of peritoneal and splenic macrophages to the cytopathic action of salmonellae.     

Experimental effect of lysozyme on macrophage metabolism and resistance to infection

The peritoneal macrophages of mice treated with lysozyme were studied by cytochemical assay. In single and repeated doses of 0.5-5 mg/kg lysozyme induced an increase in macrophage metabolism. This was evident from an increased activity of succinate dehydrogenase, NADP X N-DH and the enzymes catalyzing glycolysis typical of these cells (lactate dehydrogenase and alpha-glycerophosphate). The changes in the activity of the enzymatic systems were most pronounced in minute and less mature macrophages after repeated administrations of the drugs. In a dose of 50 mg/kg lysozyme somewhat decreased the activity of a number of the enzymes. In the doses optimal for the macrophage activity lysozyme had a low effect on the infection resistance and slightly increased the cephotaxim efficiency in experimental staphylococcal infection. This may be mainly due to the immunomodulating effect of lysozyme and its low effect on the large macrophages having the bactericidal effect.     

The effect of alpha/beta and gamma interferon preparations on the functional activity of macrophages in experimental staphylococcal infections

The preparations of alpha/beta- and gamma-interferons have been shown to stimulate the functional activity characteristics of mouse macrophages (phagocytosis, spreading, contacts with lymphocytes, bactericidal properties) obtained from the peritoneal exudate of intact animals and those infected with staphylococci. The immunomodulating action of gamma-interferon is more pronounced than that of the preparation of alpha/beta-interferon.     

Steroids mediate resistance to the bactericidal effect of phosphatidylcholines against Helicobacter pylori

Helicobacter pylori assimilates various steroids as membrane lipid components, but it can also survive in the absence of steroids. It thus remains to be clarified as to why the organism relies on steroid physiologically. In this study, we have found that phosphatidylcholine carrying a linoleic acid molecule or arachidonic acid molecule has the potential to kill steroid-free H. pylori . The bactericidal action of phosphatidylcholines against H. pylori was due to the lytic activity of the phosphatidylcholines themselves and not due to the lytic activity of the unsaturated fatty acids or lyso-phosphatidylcholine resulting from the hydrolysis of the phosphatidylcholines. In contrast to the steroid-free H. pylori , the organism that absorbed and glucosylated free cholesterol was unaffected by the bactericidal action of the phosphatidylcholines. Similarly, H. pylori that absorbed estrone without glucosylating it also resisted the bactericidal action of the phosphatidylcholines. The steroids absorbed by H. pylori existed in both the outer and inner membranes, while the glucosyl-steroids produced via the steroid absorption were localized in the outer membrane rather than in the inner membrane. These results indicate that H. pylori absorbs the steroids to reinforce the membrane lipid barrier and thereby expresses resistance to the bacteriolytic action of hydrophobic compounds such as phosphatidylcholine.     

Dysfunctional CFTR alters the bactericidal activity of human macrophages against Pseudomonas aeruginosa

Chronic inflammation of the lung, as a consequence of persistent bacterial infections by several opportunistic pathogens represents the main cause of mortality and morbidity in cystic fibrosis (CF) patients. Mechanisms leading to increased susceptibility to bacterial infections in CF are not completely known, although the involvement of cystic fibrosis transmembrane conductance regulator (CFTR) in microbicidal functions of macrophages is emerging. Tissue macrophages differentiate in situ from infiltrating monocytes, additionally, mature macrophages from different tissues, although having a number of common activities, exhibit variation in some molecular and cellular functions. In order to highlight possible intrinsic macrophage defects due to CFTR dysfunction, we have focused our attention on in vitro differentiated macrophages from human peripheral blood monocytes. Here we report on the contribution of CFTR in the bactericidal activity against Pseudomonas aeruginosa of monocyte derived human macrophages. At first, by real time PCR, immunofluorescence and patch clamp recordings we demonstrated that CFTR is expressed and is mainly localized to surface plasma membranes of human monocyte derived macrophages (MDM) where it acts as a cAMP-dependent chloride channel. Next, we evaluated the bactericidal activity of P. aeruginosa infected macrophages from healthy donors and CF patients by antibiotic protection assays. Our results demonstrate that control and CF macrophages do not differ in the phagocytic activity when infected with P. aeruginosa. Rather, although a reduction of intracellular live bacteria was detected in both non-CF and CF cells, the percentage of surviving bacteria was significantly higher in CF cells. These findings further support the role of CFTR in the fundamental functions of innate immune cells including eradication of bacterial infections by macrophages.     

Combined Action of Carbenicillin and Gentamicin on Pseudomonas aeruginosa In Vitro

The minimal inhibitory and minimal bactericidal concentrations of carbenicillin and gentamicin were determined for 10 strains of Pseudomonas aeruginosa isolated from the urinary tract. Combinations of the two drugs were tested for possible enhanced activity. Such enhancement was demonstrated in the inhibitory activity of combinations for eight strains. Striking bactericidal activity against five strains was achieved by the combination, whereas neither drug alone in low dosage was capable of bactericidal action. The possible mode of action and the possible merit of pursuing these preliminary findings are discussed.     

Thein vitro phagocytic activity of guinea-pig macrophages toSalmonella typhi andSalmonella enteritidis

The engulfing, bactericidal and degrading activities toSalmonella typhi, strain ty2-4446 and 0-901 and toSalmonella enteritidis of guinea pig macrophages obtained from peritoneal exudate, spleen and bone marrow that were cultivated for 2–7 days, were studied. The phagocytic activity was expressed as a total number of phagocytosed microbes and the number of viable bacteria, released from mechanically disrupted macrophages. The ratio of phagocytosed bacteria to the original number of bacteria that were introduced to macrophage cultures, were evaluated in per cents. No significant difference in phagocytic activity was found between macrophages submitted to thein vitro cultivation and macrophages freshly isolated from the organism. Profound variations in phagocytic activity of cells were found which were partially dependent on the dose of microbes employed for the infection of cultures. Furthermore, both the engulfing and bactericidal activity of peritoneal macrophages toSalmonella typhi were found to be higher than in bone morrow macrophages.Salmonella typhi 0-901 microbes were phagocytosed by macrophages from bone marrow and peritoneal exudate much better thanSalmonella typhi ty2. In addition, a significant delay in bactericidal activity toSalmonella typhi ty2 of bone marrow macrophages in comparison to peritoneal macrophages was observed. The spleen macrophages possessed better phagocytic and killing activity toSalmonella enteritidis than bone marrow macrophages. A striking difference was found as regards the intracellular growth ofSalmonella typhi andSalmonella gertneri: no multiplication ofSalmonella typhi within the peritoneal and bone marrow macrophages was observed during the 3–5 h cultivation, whereas on the other hand,Salmonella gertneri started to grow intracellularly within the 5 h cultivation in the bone marrow macrophages.     

Metabolic Activity of Macrophages Infected with Hantavirus,an Agent of Hemorrhagic Fever with Renal Syndrome

Monocytes/macrophages are thought to play an important role in pathogenesis of viral infections. These cells are involved in distribution and persistence of viruses in the organism and also influence the regulation of immune reactions. The functional and enzymatic activities of macrophages infected with an agent of hemorrhagic fever with renal syndrome were analyzed for the first time. This disease is caused by a virus of the Hantavirus genus, the Bunyaviridae family. Activities of ectoenzymes 5 -nucleotidase and ATPase of the plasma membrane of the hantavirus-infected macrophages decreased along with the antigen accumulation in the infected cells. The contact of phagocytes with hantavirus resulted in activation in the cells of the oxygen-dependent metabolism and NO-synthase. The NO-synthase-dependent system of the infected macrophages was activated earlier than their oxygen-dependent system. The intracellular contents of acid and alkaline phosphatases increased within the first hours after the infection. The bactericidal activity of the hantavirus-infected macrophages relatively to Staphylococcus aureus increased during the specific antigen accumulation in the phagocytes. Thus, the infection of macrophages with hantavirus was associated with intracellular metabolic changes.