Immunomodulating properties of salmozan and its effect on the functional activity of macrophages

The effect of salmozan on the resistance of mice to Listeria monocytogenes infection, the formation of delayed hypersensitivity (DH) to sheep red blood cells in the animals, as well as changes in some functional activity characteristics of macrophages have been studied. The study has revealed that salmozan enhances anti-infectious resistance, suppresses the dermal manifestations of DH, and decreases the level of 5'-nucleotidase in peritoneal macrophages, stimulating their phagocytic activity. The intensity of the drug action depends on the time of its administration. The most pronounced immunomodulating action and maximal changes in the function of macrophages have been registered simultaneously after the treatment of the animals with salmozan.     

Regulation of cell function in the macrophagal phagocytic system with salmozan

Salmozan has been shown to bring about changes in the phagocytic function of peritoneal macrophages, to produce a labilizing effect on the state of lysosomal membranes and to enhance the intensity of protein synthesis in peritoneal macrophages. The presence of relationship between the functional activity of the cells of the macrophagal phagocytic system, the resistance of the body to infections and the capacity for response to an antigenic stimulus has been revealed. The character of changes observed in this investigation has been found to depend on the interval between the injection of salmozan and the injection of sheep red blood cells or challenge with Listeria. The differences in the functional activity of the cells of the macrophagal phagocytic system, appearing under the influence of salmozan, can be attriduted to the action of this immunomodulating agent on various populations of peritoneal macrophages.     

Effect of phosphate esters, nucleotides and nucleosides on 5'-nucleotidase of cultured mouse macrophages.

Mouse peritoneal macrophages elicited by intraperitoneal injection of sodium caseinate exhibit low levels of ecto-5'-nucleotidase (E. C. 3.1.3.5) activity in contrast to macrophages obtained by peritoneal lavage. When elicited cells were cultured under standard conditions in the presence of serum, a 2.5-fold increase in 5'-nucleotidase activity was observed over a period of 48 hours. Addition of adenosine monophosphate to the culture medium led to an augmented (5-fold) increase in the specific activity (per unit cell protein) as well as an absolute increase (per culture plate) of 5'-nucleotidase. Other adenosine-containing compounds also had stimulatory effects. The levels of this enzyme thus appear to be regulated by the extracellular levels of adenosine nucleotides. The product of the enzymatic reaction--adenosine--when added to the medium exhibited a toxic effect on these cells--as did adenosine monophosphate. However, the former substance did not augment the increase in enzyme activity during culture. The toxic effect could be suppressed when the cells were cultured in the presence of uridine 5'-monophosphate. The latter substance also depressed the stimulation of enzyme activity due to AMP.     

Activity of adenosine metabolism enzymes and spontaneous chemiluminescence in macrophages in the tumor growth process

Adenosine deaminase and 5'-nucleotidase activities as well as chemiluminescence emission were measured in peritoneal macrophages of Syrian hamsters in the growth process of tumours with different grade of malignancy. The adenosine deaminase activity was established to decrease, while the 5'nucleotidase activity--to increase in macrophages after the subcutaneous injection of tumour cells with high level of malignancy as compared with these values in normal cells. This is accompanied by a decrease of the macrophage chemiluminescence during the whole experimental period. At the same time adenosine deaminase and 5'-nucleotidase activities as well as chemiluminescence emission in peritoneal macrophages of hamsters treated with low-malignancy cells do not differ from these values in the control group.     

Action of plant extracts on the natural immunity indices of animals

The influence of extracts from oak bark, St. John's-wort leaves and pine buds on natural immunity characteristics of mice has been studied. The injection of these extracts into mice has been found to enhance their resistance to infection with Staphylococcus aureus and Bordetella pertussis virulent cultures, to decrease the enzymatic activity of 5'-nucleotidase in the peritoneal exudate macrophages of mice and to increase the level of lysozyme in their blood. The action of these extracts has proved to depend on their dosage and the time of observation.     

Analysis of the effect of immunostimulants on the macrophages of mouse peritoneal exudates by using mathematical simulation and planning methods

Using methods of mathematical modelling and planning an analysis was made of changes in 5-nucleotidase activity in murine peritoneal macrophages after intraperitoneal administration of immunostimulants of different chemical structure and biological origin. The changes in 5-nucleotidase activity after the administration of immunostimulants were shown to exhibit a similar linear pattern.     

Macrophage activation by Lactobacillus casei in mice

Effects of Lactobacillus casei YIT 9018 (LC 9018), which has antitumor activities against allogeneic and syngeneic murine tumors, on macrophage functions were examined. By intraperitoneal (i.p.) injection of LC 9018, acid phosphatase activity and phagocytic activity of peritoneal macrophages were enhanced significantly compared with those of normal peritoneal macrophages. The phagocytic activities showed peaks 2-3 days after the LC 9018-injection. LC 9018 accelerated the phagocytic function of the reticuloendotherial system in ICR mice tested by the carbon clearance test. The cytostatic activity of peritoneal exudate cells (PEC) induced by i.p. injection of LC 9018 into C57BL/6 mice against EL4 cells was also enhanced. On the other hand, PEC induced by L. fermentum YIT 0159, which has no antitumor activity, did not have cytostatic activity. These observations showed that LC 9018 was able to activate macrophages in mice.     

Sensitivity to mouse toxin and level of macrophage 5-nucleotidase activity]

It is shown that immunomodulator of bacterial origin salmozan causes alternations of sensitivity to mouse toxin in mice CBA. A correlation exists between the sensitivity to mouse toxin and the level of 5-nucleotidase of peritoneal exudate macrophages.     

Carnosine as a stimulator of cytotoxic and phagocytic function of peritoneal macrophages]

Biochemical changes in peritoneal macrophages and their relatedness to the cytostatic and phagocytotic function in C3HA mice injected with a single intraperitoneal dose of 0.45 mM carnosine and 4-methyluracil or stimulated with peptone have been studied. During the first 24 hours after injection both carnosine and 4-methyluracil increase the activity of adenosine deaminase and purine nucleoside phosphorylase, the key enzymes of purine catabolism which is the main source of O2-. radicals in macrophages. In carnosine-stimulated macrophages the activity of membrane 5'-AMP nucleotidase decreases on days 1-3 after injection which points to alleviation of adenosine-induced inhibition as well as to macrophage activation. Carnosine increases the cytostatic and phagocytotic activities of macrophage coupled to O2-. production. The mechanism of the stimulating effect of carnosine on macrophages seems to consist in the dipeptide interaction with specific receptors localized on the plasma membrane of macrophagal cells.     

Effect of mytilan, a polysaccharide from Crenomytilus grayanus, on the functional activity of peritoneal macrophages

The studies showed that mytilan, a polysaccharide from Crenomytilus grayanus had a marked activating effect on macrophages evident from respective morphological changes and increased metabolic and functional activity of the macrophages. Exposure to mytilan resulted in an increase in the number of the macrophages and their size (optic microscopy), changes in their surface (scanning microscopy) and ultrastructural reconstruction of the cells (light microscopy). It was shown that subcutaneous and intraperitoneal administration of mytilan defined a decrease in the activity of 5'-nucleotidase in the macrophages of the peritoneal exudate from ++noninbred mice and mice CBA. Moreover, under the influence of mytilan the macrophage phagocytic activity against E. coli, S. aureus, P. aeruginosa and P. vulgaris increased in vivo and in vitro.     

Modulation of murine macrophage function by N-acetylcysteine in a model of endotoxic shock

In previous studies we have observed changes in several functions of peritoneal macrophages from BALB/c mice with irreversible endotoxic shock caused by intraperitoneal injection of E. coli lipopolysaccharide (LPS) (100 mg/kg), which were associated with a high production of superoxide anion. Since antioxidants, such as N-acetylcysteine (NAC), are free radical scavengers that improve the immune response, in the present work we have studied different functions of peritoneal macrophages from BALB/c mice suffering the endotoxic shock above indicated and administered N-acetylcysteine (150 mg/kg i.p.) at 30 minutes after LPS injection. In the peritoneal macrophages obtained at 2, 4, 12 and 24 h after LPS injection, the following functions were studied: adherence to substrate, mobility, ingestion of particles, and production of superoxide anion and tumour necrosis factor (TNF alpha). The increase in adherence, ingestion and superoxide anion and TNF alpha production shown by macrophages from animals with endotoxic shock was counteracted by NAC injection. Moreover, the survival time of mice with endotoxic shock was increased in the presence of NAC. These data suggest that NAC, administered intraperitoneally, may be useful for the treatment of irreversible endotoxic shock by modulation of the function of macrophages with decreased superoxide anion and TNF alpha production and concomitant increase of survival time.     

Differences in the receptor apparatus of resident and meat-peptone broth-stimulated macrophage populations detectable under the action of salmozan

The degree of expression of lectin-like receptors in resident (RPM) and peptone-elicited peritoneal macrophages (PE-PM) using salmozan as polysaccharide was investigated. Obtained results allow to suppose that salmozan screens lectin-like receptors in macrophages and SRBC. The salmozan screening of lectin-like receptors in macrophages and SRBC resulted in decrease of SRBC uptake level in RPM-subpopulation but did not alter it in PE-PM-subpopulation. The same effect was occurred with the intensified erythrocyte loading of this macrophage subpopulations. It is suggested that one of the functional differences of studied macrophage subpopulation consists in the largest expression of lectin-like receptors in RPM-subpopulation.     

The effect of muramyl dipeptide analog, GMPD, incorporated into liposomes of various composition on the functional activity of macrophages

Liposomes of different composition and N-acetylglucosaminyl-N-acetylmuramyl-L-alanyl-D-isoglutamine (GMPD) encapsulated in them are studied for their effect on the functional activity of macrophages by means of determining the 5'-nucleotidase and adenosine deaminase activity in the in vivo experiments. It is shown that both liposomes and GMDP encapsulated in them increase the activity of adenoside deaminase and decrease that of 5'-nucleotidase. This evidences for a change in the adenosine metabolism in the alveolar and peritoneal macrophages and an increase in the functional activity of cells which resulted from that rise. The manifestation of the process depends both on the lipid composition of liposomes and their charge. The observed increase in the functional activity of the alveolar macrophages under the effect of liposomes and GMDP encapsulated in them correlates with inhibition of the lung metastases development in mice.     

Studies of latent cytomegalovirus infection: the macrophage as a virus-harboring cell

During chronic infection of mice with mouse cytomegalovirus (MCMV), the virus was isolated from various tissues by cocultivation with allogeneic mouse embryonic fibroblasts (MEF). Infectious virus was recovered from over 15% of the pancreases, salivary glands, kidneys, lacrimal glands, and spleens. When activated macrophages were obtained by intraperitoneal injection of peptone into mice infected 3 months earlier, they harbored MCMV. Macrophages or lymphocytes were infected with MCMV in vitro and injected into normal mice intravenously. The peritoneal cavities of these mice were then stimulated by peptone injection 3 months after the transfer, and peritoneal or splenic macrophages and lymphocytes were cocultured with allogeneic MEF. MCMV was recovered from the peritoneal and splenic macrophages and not from the lymphocytes.     

The functional activity of the macrophages in experimental intestinal dysbacteriosis in rats

The study of the functional activity of macrophages in the peritoneal exudate and spleen of Sprague-Dawley rats was made under the conditions of dysbacteriosis caused by the administration of antibiotics (gentamicin and phenoxymethylpenicillin) and short-term food deprivation. The complex study of the key functions of macrophages, carried out on the same pool of cells, showed that under the conditions of antibiotic therapy the enhanced function of ingestion was accompanied by a decrease in all other investigated functional parameters of peritoneal macrophages: the intensity of protein synthesis, oxidation metabolism, the activity of lysosomal apparatus, 5'-nucleotide activity. In contrast to changes observed in macrophages after the administration of antibiotics, short-term food deprivation induced increased functional activity as shown by most tests used in this investigation (such as tests for the ingestion of sheep red blood cells, 51Cr, nitro blue tetrazolium, acridine orange, 5'-nucleotidase activity), but did not affect the intensity of protein synthesis in macrophages. The comparison of these results with the data obtained by the authors after the administration of an immunomodulating agent made it possible to regard short-term food deprivation, judging by its effect on the functional activity on macrophages, as an immunostimulating action.     

The action of pertussis vaccines and GMDP on the change in the enzymatic activity of macrophages from peritoneal exudate

The influence of whole-cell and acellular pertussis vaccines, introduced both alone and in combination with N-acetylglucosaminylmuramyl-2-alanine-D-isoglutamine (GMDP) on the activity of two enzymes of peritoneal exudate macrophages (5'-nucleotidase and Na+K(+)-adenosine triphosphatase) was studied. The study revealed that both pertussis vaccines exhibited immunomodulating properties, these properties being most pronounced in whole-cell pertussis vaccine. The use of GMDP in combination with pertussis vaccines led to changes in the enzymatic activity of peritoneal exudate macrophages, which was indicative of a decrease in the immunomodulating action of pertussis preparations.     

Induction of natural killer cell activity in mice by injection of indomethacin

The natural killer (NK) cell system of mice in the peritoneal cavity is of very low to undetectable activity, and testing peritoneal NK cells is a useful model to study the influence of activating substances upon local injection. Injection of indomethacin at doses of 100-400 micrograms/mouse caused a marked activation of NK cell activity which was maximal at 3 days and lasted for a total of 6 days. A similar albeit less marked effect was observed with other cyclooxygenase inhibitors such as aspirin. Prostaglandin E2 reversed the activation of NK cells induced by injection of indomethacin. The cellular count of the peritoneal population was 2-fold elevated after indomethacin injection but the percentage of macrophages in the washed-out cell population was decreased from 60% (controls) to around 20%. The NK cell nature of the effector cells activated by indomethacin was substantiated by the finding that previous injection of anti-asialo GM1 antibody prevented activation. Interferon could not be detected in the peritoneal wash fluid after injection of indomethacin, suggesting interferon-independent activation. However, the possibility of small interferon quantities being locally produced could not be excluded. In further experiments we found after intraperitoneal injection of indomethacin not only cells that killed YAC-1 targets in a 4-hour assay but also killer cells that were insensitive to anti-asialo GM1 and killed P815 cells in an 18-hour assay. We assumed that these were macrophages and have done further experiments with in vitro grown bone-marrow-derived macrophages. These could be activated for killing of P815 targets by the addition of indomethacin, but (to a lesser degree) also for killing of YAC-1 lymphoma cells.     

Change in activity of enzymes for purine metabolism and RNA and DNA biosynthesis in macrophages, reflecting impairment of their functions in neoplastic growth

The changes in the biochemical parameters of peritoneal macrophages and their coupling to the secretory and phagocytic functions in CH3A mice during the growth of the reinoculated solid hepatoma 22a were studied. The DNA and RNA synthesis during the active tumour growth was more intense than that in resident macrophages. The activity of uridine kinase increased up to 156.0 +/- 12.0 nmol/hour/10(8) but was absent in resident macrophages. This was accompanied by a 7.2-fold increase of interleukin-1 synthesis as determined by the [3H]thymidine incorporation into thymocyte DNA in response to concanavalin A administration to C3H mice. Similar changes were observed in peptone-stimulated macrophages. A specific feature of macrophages from tumour-bearing mice was the impairment of activity of purine exchange enzymes and the efficiency of phagocytosis that were unobserved in peptone-stimulated macrophages. The activity of adenosine deaminase and purine nucleoside phosphorylase was inhibited as a result of their preincubation with zymosan, a phagocytosis-stimulating agent. This was accompanied by a significant decrease of the first chemiluminescence peak resulting from disturbances in Fc-reception. Macrophages of tumour-bearing animals possessed an increased 2.2-fold activity of membrane-bound AMP 5'-nucleotidase concomitant with the lack or decrease of the amplitude of the second chemiluminescence peak reflecting the disturbances in digestion resulting from phagocytosis.     

Functional macrophage activity in infection with virulent and avirulent strains of the dysentery microbe

The effect of S. flexneri virulent and avirulent (vaccine) strains 2a on the cytoplasmic membrane of mouse macrophages has been studied by evaluating the action of these bacteria on the activity of 5-nucleotidase. The dynamics of the activity of 5-nucleotidase after the introduction of both virulent and avirulent strains has a phasic character with alternating rises and falls in the activity of this enzyme in comparison with the control. S. flexneri vaccine strain produces mainly a stimulating effect on the functional activity of peritoneal macrophages in mice, which is confirmed by a decrease in the activity of 5-nucleotidase; on the contrary, S. flexneri virulent strain- has mainly an inhibiting effect on the functional activity of peritoneal macrophages, which is confirmed by an increase in the activity of 5-nucleotidase in these cells. The comparative study of changes in the activity of 5-nucleotidase, following the introduction of S. flexneri, in mice, previously immunized with smallpox vaccine, and in intact mice has shown that the use of animals immunized with smallpox vaccine in the study of metabolic characteristics may lead to distortions in the results of the experiment.     

Properdin deficiency in murine models of nonseptic shock

Hereditary properdin deficiency is linked to susceptibility to meningococcal disease (Neisseria meningitidis serotypes Y and W-135) with high mortality. Its relative contribution toward the outcome of nonseptic shock has not been investigated. Using properdin-deficient C57BL/6 mice and their littermates, this study examines their survival of zymosan-induced and LPS-induced shock. Properdin-deficient mice were more resistant to zymosan shock compared with wild-type mice, which showed greater impairment of end-organ function 24 h after zymosan injection, higher TNF-alpha production by alveolar and peritoneal macrophages, higher TNF-alpha, and, inversely, lower IL-10 levels in peritoneal lavage and circulation and higher plasma C5a levels. Properdin-deficient mice showed significantly higher mortality in LPS shock, elevated TNF-alpha, and, inversely, reduced IL-10 production by peritoneal macrophages as well as lower plasma C5a levels compared with wild-type littermates. NO production by peritoneal macrophages and plasma alpha1-antitrypsin levels at 24 h after the injection of LPS or zymosan were decreased in properdin-deficient mice in both models, and fewer histopathologic changes in liver were observed in properdin-deficient animals. This study provides evidence that properdin deficiency attenuates zymosan-induced shock and exacerbates LPS-induced shock.