IL-12 and IL-18 synergistically induce the bactericidal activity of murine peritoneal cells against M. leprae.

We examined the effect of IL-12 and IL-18 on bactericidal activities of mouse peritoneal cell (PC) against Mycobacterium leprae (M. leprae). We demonstrated that IL-12 and IL-18 synergistically induced the NO-dependent bactericidal activity of PC by stimulating Natural Killer (NK) cells and T-cells through IFN-gamma production. IL-12 and IL-18 induced host cell death through NK-cells and T-cells. Therefore. IL-12 and IL-18 play an important role on direct killing of intracellular M. leprae and on indirect killing of them through inducing host cell death.

[Monokine production by mouse peritoneal macrophages after phagocytosis of mycobacteria].

It is well known that monokines, IL-1 (Interleukin-1) and TNF (Tumor Necrosis Factor), are produced by macrophages after stimulated with various agents. These cytokines are involved in various aspects of the inflammatory process and immunological response in addition to their original activities to proliferate T lymphocytes and causing tumor necrosis, respectively. Recently, there have been reported that IL-1 and TNF also play an important role in mycobacterial infections such as granuloma formation. In the present study, IL-1 and TNF productions were observed by mouse peritoneal exudate and resident macrophages after incubation with heat-killed M. lepraemurium and M. avium in vitro. The production was enhanced by phagocytosis of these mycobacteria in a dose dependent manner, and the time course of the production was maximum within 24 hr after phagocytosis of these mycobacteria. It was also shown of morphological changes and enhanced glucose consumption in media by these macrophages. Above results suggest that phagocytosis of mycobacteria by macrophages leads to monokine production, which would not only causes well known immunological reactions but also makes characteristic phenomena to be observed in mycobacterial infections.

Effect of anti-leprosy drugs on superoxide anion production by rat peritoneal macrophage with special reference to light exposed clofazimine.

The present study describes the in vitro effect of anti-leprosy drugs on superoxide anion (O2-) production by rat resident peritoneal macrophages. Of the three drugs tested i.e. clofazimine, rifampicin and dapsone, the first was most effective in increasing O2- production in a dose dependent manner, while rifampicin had some stimulatory effect and dapsone exhibited minimal action. Furthermore, when clofazimine and dapsone were added together it was observed that the increase of O2- production by macrophages due to clofazimine was not significantly altered by the addition of dapsone. Moreover, it was found that killed Mycobacterium leprae could induce a lesser amount of O2- production in comparison to that of Staphylococcus aureus and the enhancement of O2- release due to clofazimine was stimulus dependent. This increase of O2- release after addition of clofazimine was inhibited by the addition of p-bromophenacyl bromide. Another interesting finding was that the enhancement of O2- production by clofazimine gradually decreased as clofazimine was exposed to light for days. On further investigation it was found that ultraviolet, NMR, infrared and mass spectra of the light unexposed and exposed drug were similar, but the diffusion current of the polarogram of light exposed drug was remarkably more than that observed in light unexposed drug, indicating, thereby, a possible increase in the electron accepting capacity of the light reacted molecule. As far as we know this is the first report describing the effect of light exposed clofazimine on the respiratory burst activity of macrophages.

Variation in immunogenicity of mycobacteria: role of antigen-presenting cells.

The antigen-presenting efficiency of peritoneal cells and irradiated spleen cells was compared using Mycobacterium tuberculosis- and M. vaccae-primed T cells and corresponding sonicates as antigens in an in vitro lymphocyte transformation test. The presentation efficiency of irradiated spleen cells was reasonably good for both antigens. However, with peritoneal cells as the antigen-presenting cells, the proliferative response against only M. tuberculosis sonicate was good. Proliferation of M. vaccae-primed T cells was very poor when the antigen was presented by peritoneal cells. Poly I:poly C treatment of mice prior to harvesting the peritoneal cells resulted in distinct improvement in their efficiency to present M. vaccae sonicate; maximal proliferative response was obtained with peritoneal cells from mice receiving two and three doses of poly I:poly C 24 hr apart. Even paraformaldehyde-fixed peritoneal cells from poly I:poly C-treated mice gave an efficient M. vaccae-specific stimulation to primed T cells. Based on these data, it was concluded that failure of mice to respond to M. vaccae by intraperitoneal immunization is the result of the poor efficiency of presentation of M. vaccae antigen.

[The modelling of chronic infection (experimental leprosy) against a background of macrophagal immunodeficiency]. / Modelirovanie khronicheskoi infektsii (éksperimental'noi lepry) na fone makrofagal'nogo immunodefitsita.

The dynamics of mycobacterial multiplication was followed in mice with intraplantar leprosy infection and preinduced macrophage insufficiency. The characteristics of Shepard's model appeared to be similar to those of the method proposed by the authors including the susceptibility to the main antileprosy drugs. Peritoneal macrophages were cytochemically studied in the process of development of mononuclear phagocyte deficiency and experimental leprosy. It was concluded that the method proposed preserving all the merits of Shepard's model should allow one to shorten significantly the duration of testing potential drugs for their antileprosy activity.

[Phospholipid-deacylating activities of the mouse peritoneal macrophages during phagocytosis].

In the macrophages (M phi) obtained from mouse peritoneal exudates, five kinds of phospholipid-deacylating activities were detected using phosphatidylethanolamine (PE) and phosphatidylcholine (PC) labeled with [1-14C]oleic acid either in 1- or 2- position and 1- [1-14C]oleoyl-lysoPE, as substrates. Two types of phospholipipase A1 with pH optima of 4 to 6 and 8, respectively, and two types of phospholipase A2 activities with pH optima of 4 to 5 and 8, respectively, were identified. A detected lysophospholipase activity exhibited a broad pH optimum between 4 and 8. Both types of the phospholipase A1 and A2 of M phi hydrolyzed PE more than PC. Exogenously added Ca2+ did not increase the enzymatic activities. A comparison was made of three kinds of the M phi-phospholipid deacylating activities at pH8, after challenging the M phi with Mycobacterium lepraemurium, Escherichia coli, zymosan, or latex beads for 17 hours at 37 degrees C. The bacteria used to the phagocytosis were autoclaved. When the M phi were challenged with M. lepraemurium, the phospholipase A1, A2 and lysophospholipase activities were stimulated by about 160%, 150% and 140%, respectively. However, when challenged with E. coli, the phospholipase A1 activity remarkably decreased by about a third, although the phospholipase A2 activity was stimulated by about 150% that is similar to the challenge with M. lepraemurium. An inflammatory substance, zymosan seemed an effective inducer of the phospholipase A2, the enzymatic activity was remarkably stimulated by 260%, when challenged with 200 micrograms of zymosan. The increase in phospholipase A2 activity of the M phi pretreated with the bacteria or zymosan seems to result in an increase in the hydrolysis of arachidonic acid from the M phi-phospholipids to synthesize its inflammatory oxygenated metabolites. The lysophospholipase activity was not stimulated by the substances used to challenge the M phi, except for M. lepraemurium. No significant increase in three kinds of phospholipid-deacylating activities was observed after challenging the M phi with latex beads. It was considered from the above results that the M phi-phospholipid-deacylating activities at pH8 might be affected by sort of the ingested substances.

Accessory cell function of cells isolated from Mycobacterium leprae-induced granulomas.

The large cells from Mycobacterium leprae-induced granulomas in guinea pig lymph nodes were separated by Percoll discontinuous density gradient centrifugation and on a fluorescence-activated cell sorter (FACS) using cross-reacting monoclonal antibody to human MHC Class II antigens. Large Percoll-separated cells (83% Class II antigen positive and 52% macrophage-specific antigen positive) and FACS-separated cells are able to act as antigen-presenting cells for T-cell proliferation to PPD. In previous studies, macrophage antigen-positive cells consistently failed to act as accessory cells. This indicates that there is a population of accessory cells which are macrophage antigen negative and MHC Class II antigen positive present in these M. leprae-induced granulomas.