In Vitro Study of the Expression of CD1, CD14, CD25, CD30, CD35, CD95 Receptors by Macrophages of Mice Infected with Mycobacterium tuberculosis.

In cultures of peritoneal macrophages (MP) of male BALB/c mice infected with Mycobacterium tuberculosis from the BCG vaccine, the expression of CD1, CD14, CD25, CD30, CD35, and CD95 receptors was studied in vitro 3 months after infection. In MP cultures from intact and infected mice, mononuclear MP predominated (96 and 92%, respectively). Bi- and trinuclear MP in MP cultures from control and infected mice constituted 4 and 8.3% of all MP, respectively. In the cultures of both groups, no obvious correlations between the number of MP expressing CD-receptors and number of nuclei in these cells were found, but the expression of CD14 receptor was more often noted. In cultures from infected animals, hypertrophied MP and enhanced (by several times) expression of all CD-receptors were observed. The increase in the expression of CD-receptor can be determined by activation of plastic processes in hypertrophied MP (in epithelioid and in numerically insignificant polynuclear MP), which is due to the phenomenon of prolonged M. tuberculosis persistence in the vacuolar apparatus of these cells.

Modifying Effects of Nanosized Diamonds on Hydrolytic Potential of Macrophages In Vitro.

The effects of nanosized particles (4-6 nm) of synthetic diamonds on mouse macrophage expression and secretion of lysosomal cathepsins (B and D) and MMP-1 and MMP-9 were studied in vitro. Culturing of peritoneal macrophages in medium with diamond nanoparticles led to an increase in the counts of macrophages expressing the above enzymes and to stimulation of their secretion. However, the manifestations of these effects varied significantly for various enzymes. The data indicate modulation of macrophage functions by nanodiamonds. These results help better understand the possible role of the "corpuscular" xenobiotic factors in the pathogenesis of diseases associated with macrophage capturing of these factors irrespective of their chemical "activity".

In vitro effects of nanosized diamond particles on macrophages.

The effects of synthetic diamond nanoparticles (4-6 nm) on mouse macrophage biotropism and biocompatibility and the modulation of the macrophage functions (expression of IL-1α, TNF-α, GM-CSF, bFGF, and TGF-ß) by nanoparticles in different concentrations were studied in vitro during exposure of different duration. Macrophage endocytosis of nanodiamonds increased with increasing the concentration of nanoparticles in culture and incubation time. Nanodiamonds exhibited high biotropism and biocompatibility towards macrophages; in doses of 10-20 µg/ml, they induced expression of GM-CSF and TGF-ß, inhibited expression of bFGF, and did not stimulate IL-1α and TNF-α. These data indicate that nanodiamond capture by macrophages in the studied experimental model led to modulation of the functional status of macrophages that determine their capacity to stimulate reparative processes without increasing proinflammatory and profibrogenic status.

In vitro study of phenotypical characteristics of BCG granuloma macrophages over the course of granuloma development.

Structural and immunophenotypical characteristics of macrophages, associated with their polarization in the M1 and M2 directions of differentiation and activation, were studied in different morphological types of BCG granulomas by the expression of GM-CSF, IFN-γ, and FGFb cytokines and CD36 and CD16/32 differentiation clusters. The proportion of IFN-γ and FGFb macrophage subpopulations changed over the course of granuloma formation, which led to accumulation of FGFb macrophages in the granulomas. These data indicated that the formation of macrophage granulomas and their subsequent transformation into epithelioid-cell granulomas were associated with dynamic quantitative changes in the subpopulations of macrophages with the morphofunctional characteristics of M1 and M2 phenotypes, determining the antibacterial and destructive potential of granulomas. These data are useful for understanding the contribution of functional polarization of macrophages to the pathogenesis of tuberculous infection and, presumably, its complications.

Study of macrophages in BCG granulomas in different compartments of the mononuclear phagocyte system.

We studied in vitro morphological and functional properties of macrophages associated with their M1 and M2 polarization in different mononuclear phagocyte compartments during BCG-induced granuloma formation, namely expression patterns of cytokines IL-1α, GM-CSF, TNF-α, and clusters of differentiation CD36 and CD16/32. We showed the mosaic pattern of macrophage polarization in BCG granulomatosis manifested by simultaneous formation of different macrophage subpopulations with M1 and M2 phenotypes in the population of mononuclear phagocytes of BCG granulomas and various compartments of the mononuclear phagocyte system. These data clarify the role of the functional polarization of macrophages in the pathogenesis of tuberculosis infection.

Dynamics of structural transformations of BCG granulomas and expression of TNF-α and granulocyte-macrophage CSF by macrophages in vitro.

The spleens were isolated from mice at different times after BCG infection and BCG granulomas were explanted and cultured in vitro. Cell migration, chemoattractant potential, and expression of TNF-α and granulocyte-macrophage CSF (GM-CSF) by macrophages migrated from granulomas were evaluated in granulomas. The number of macrophages able to migrate; migrating out of granulomas, expressing TNF-α and GM-CSF decreased with increasing the time after infection. The number of cells in "dissociating" granulomas correlates with the number of macrophages containing live BCG mycobacteria in the cytoplasm.

Phagocytosis of hybrid molecular nanosomal compositions containing oxidized dextrans conjugated with isonicotinic acid hydrazide by macrophages.

We studied phagocytic activity of macrophages towards hybrid molecular nanosomal compositions consisting of 150-800-nm nanoliposomes containing oxidized dextrans with a molecular weight of 35 and 60 kDa obtained by chemical ("permanganate") and radiochemical oxidation of dextran conjugated with isonicotinic acid hydrazide (dextrazides, intracellular prolonged antituberculous drugs). Phagocytic activity of macrophages towards hybrid molecular nanosomal compositions containing dextrazides obtained by chemical oxidation of dextrans is higher than activity towards hybrid molecular nanosomal compositions containing dextrazides prepared by radiochemical oxidation and depends on the size of hybrid molecular nanosomal compositions and molecular weight of oxidized dextrans.

Phagocytic activity of macrophages against liposomes with conjugates of oxidized dextrans and isonicotinic acid hydrazide during modeling of phagocytosis disturbances in vitro.

We studied phagocytic activity of macrophages against molecular-liposome hybrid compositions consisting of liposomes (diameter 200-450 nm) containing oxidized dextrans with a molecular weight of 35 or 60 kDa conjugated with the basic antituberculosis preparation isonicotinic acid hydrazide (dextrazides) during modeling of various disturbances of endocytosis function of phagocytic cells in vitro. Preincubation of macrophages with trypsin, colchicine, or sodium azide did not change the parameters of adhesion of molecular-liposome hybrid compositions to macrophages. It was found that preincubation of cells with colchicine or sodium azide reduced parameters of phagocytosis of the molecular-liposome hybrid compositions; this reduction did not depend on the molecular weight of dextrans entering the composition of the molecular-liposome hybrid compositions.

Effects of molecular liposomal hybrid compositions with oxidized dextrans and isonicotinic acid hydrazide on production of granulocytic macrophage colony-stimulating factor by macrophages.

The effects of molecular liposomal hybrid compositions consisting of liposomes (200-450 nm) containing oxidized dextrans (dextranals; 35-60 kDa) conjugated with isonicotinic acid hydrazide (dextrazides), their components, and native dextrans on the production of granulocytic macrophage CSF by peritoneal macrophages were studied in vitro. Dextranals proved to be more potent inductors of granulocytic macrophage CSF than native dextrans. Conjugation of nicotinic acid hydrazide with dextranals did not modify their capacity to stimulate the production of granulocytic macrophage CSF. Liposomes in the molecular liposomal hybrid compositions did not attenuate the dextrazide capacity to stimulate the production of granulocytic macrophage CSF. Molecular liposomal compositions containing 60 kDa dextrazide exhibited the most potent stimulatory effect on macrophage production of granulocytic macrophage CSF.

In vitro effect of oxidized dextrans on peritoneal cells.

We studied the dependence of in vitro dextran biocompatibility on the method of oxidation of 35-kDa dextran. The biocompatibility of dextran oxidized with potassium permanganate was higher compared to that obtained by radiochemical oxidation. It was related to the formation of peroxide compounds during radiochemical oxidation.

Comparative study of the in vitro effect of nanoliposomes with oxidized dextrans on peritoneal cells.

We studied the in vitro effect of hybrid molecular-nanosomal biocompatible compositions on cultured peritoneal cells. The compositions consisted of oxidized dextrans with a mean molecular weight of 35 and 60 kDa, which were obtained by chemical and radiochemical oxidation of dextran. Hybrid nanoliposomal compositions of chemically oxidized dextran (permanganate method) had greater biocompatibility and tropic activity to macrophages compared to nanoliposomes of radiochemically oxidized dextran.