Esfandiar Ghadirian

The requirement for gamma Interferon in resistance of mice to experimental tularemia

The role of gamma interferon (IFN-gamma) in the host response to experimental tularemia was evaluated in a murine model. C57BL/6 strain mice were given a series of daily intravenous injections of 10(6) units (U) recombinant murine IFN-gamma prior to infection with Francisella tularensis LVS. Three days later, the number of bacteria in the tissues of IFN-gamma-treated mice was found to be less than that in control mice by a factor of 10-20. The effect of IFN-gamma on anti-tularemic resistance was dependent upon the administered dose, with as little as 10(4) U/mouse/day inducing a significant level of enhanced resistance. IFN-gamma was also effective in enhancing resistance to tularemia in the A/J mouse strain which, in comparison with the C57BL/6 strain, is more susceptible to infection. When C57BL/6 mice were treated with a monoclonal antibody directed against murine IFN-gamma, the number of Francisella recovered from their tissues 6 days following infection was increased by as much as 15 times, in comparison with control mice. The results of these experiments clearly indicate that the resolution of experimental murine tularemia is dependent, at least in part, on the participation of IFN-gamma.

Role of mononuclear phagocytes in elimination of Plasmodium chabaudi AS infection

Summary The role of mononuclear phagocytes in acquired immunity resulting in the intraerythrocytic destruction and elimination of malarial parasites was investigated in the murine model of infection with Plasmodium chabaudi AS. Mice were treated 1 day before or 6 days after infection with agents which either result in augmentation or activation of the non–specific, microbicidal effector function of mononuclear phagocytes or in depletion of cells of this lineage. To examine the effect of agents which activate mononuclear phagocytes, A/J mice, which are susceptible to P. chabaudi AS and exhibit fulminant parasitaemia and death within 10 days of intraperitoneal infection with 106 P–RBC, were treated intravenously with muramyl dipeptide (MDP) or liposome–encapsulated MDP–glycerol dipalmitate (MDP–GDP). Treatment administered 1 day before infection was ineffective. Treatment on day 6 post–infection with liposome–encapsulated MDP–GDP (1 ftg) resulted in a significant decrease in parasitaemia on day 8 and survival, while treatment with free MDP (100 ^g) resulted only in a significant decrease in parasitaemia. To examine the effect of depletion of mononuclear phagocytes, C57BL/6 mice, which are resistant to P. chabaudi AS infection and eliminate the parasite by 4 weeks, were treated intravenously with 3 mg silica. Silica administered 1 day before or 6 days post–infection abrogated resistance resulting in a delay in elimination of the parasite and host mortality. Treatment on day 6 was more effective, with death by day 13 post–infection of 70% of the normally resistant C57BL/6 mice which exhibited fulminant parasitaemia levels. These results thus provide in–vivo evidence that mononuclear phagocytes play a critical role in the elimination of infection with the murine malaria species P. chabaudi AS. Furthermore, these results suggest that the time of administration of agents which alter mononuclear phagocyte function may be important in determining their effect on host antimalarial defences.

Granulocyte-macrophage colony-stimulating factor restricts growth of tubercle bacilli in human macrophages

The effect of human recombinant granulocyte-macrophage colony stimulating factor (GM-CSF) on the uptake and intracellular growth of Mycobacterium tuberculosis in human macrophages was investigated. GM-CSF was found to reduce growth of M. tuberculosis in human monocyte-derived macrophages in vitro, as compared to untreated cells. The decrease in intracellular growth of the tubercle bacilli did not depend on generation of reactive oxygen species, inasmuch as treatment with scavengers did not affect the ability of GM-CSF to restrict mycobacterial growth in macrophages.

Transforming growth factor beta (TGF-b1) plays a detrimental role in the progression of experimental Mycobacterium avium infection; in vivo and in vitro evidence

BALB/c mice were infected with 10(5) colony forming units (cfu) of Mycobacterium avium TMC 702 i.v. and the growth of the inoculum followed in the spleens of control mice. Other infected mice given weekly doses of 1 microgram of TGF-b1 or weekly doses of 2 mg of a rabbit antiserum against mouse TGF-b1 were evaluated for their resistance to M. avium TMC 702. Growth of M. avium in the spleens of mice given repeated doses of TGF-b1 (1 microgram weekly) was significantly higher than in the spleens of control mice starting at day 40 of infection. Similarly, growth of M. avium was significantly diminished (0.7 log difference at 80 days) in mice given infusions of anti-TGF-b1 (2 mg weekly). Macrophage activation status was similar in the three groups of mice, as seen by a comparable release of superoxide anion (O2-) and hydrogen peroxide (H2O2) by peritoneal macrophages of infected mice. However, TGF-b1-pulsed peritoneal macrophages were found to be more permissive for M. avium growth in vitro than control macrophage monolayers. Overall, these results suggest that TGF-b1 plays a detrimental role in the progression of experimental M. avium infections, by an unclear mechanism.

In vivo activation of macrophages by IFN‐γ to kill Entamoeba histolytica trophozoites in vitro

Summary To determine the role of interferon‐gamma (IFN‐γ) in the activation of macrophages to kill Entamoeba histolytica trophozoites in vitro, C57BL/6 mice were injected with various doses of recombinant IFN‐γ (rIFN‐γ) by either the intravenous, intraperitoneal or intramuscular routes. Mice were treated with doses of rIFN‐γ ranging from 101 to 105 units. Twenty hours later, peritoneal macrophages were harvested from the treated animals. Macrophage monolayers were prepared and their in vitro cytotoxic activity against a virulent strain of E. hystolytica (IP:0682:1) was determined. Amoebicidal activity was determined by counting the number of dead trophozoites by Trypan Blue exclusion in cultures containing macrophages and amoebic trophozoites which were incubated together for 4 h. Both intravenous and intraperitoneal treatment resulted in the recovery of macrophages from the peritoneal cavity which exhibited amoebicidal activity in vitro. Peritoneal macrophages harvested from mice that had been treated intraperitoneally or intravenously with rIFN‐γ, however, showed significantly more amoebicidal activity in comparison to macrophages harvested from animals treated intramuscularly. There was a dose dependent relationship between the concentration of rIFN‐γ used to activate macrophages in vivo and the number of dead trophozoites in vitro. In addition, these results confirm our previous observations that treatment in vitro with rIFN‐γ can activate murine peritoneal macrophages to kill amoebic trophozoites.

Activated mouse macrophages kill Entamoeba histoloytica trophozoites by releasing reactive nitrogen intermediates

Mouse macrophages activated by gamma interferon (IFN-gamma) and bacterial lipopolysaccharide (LPS) are highly cytotoxic for the enteric protozoan parasite Entamoeba histolytica. Herein, we show that this killing by activated macrophages is L-arginine dependent, inasmuch as it was blocked by exogenous arginase or NG-monomethyl-L-arginine. These two inhibitors had no effect on E. histolytica cytolytic activity against L929 fibroblasts. Also, macrophage killing of E. histolytica always correlated with nitrite presence in the supernatant fluids. Finally, it was shown that addition of excess iron or the reductant sodium dithionite to activated macrophages blocked their ability to kill E. histolytica. Overall, this suggests that killing of E. histolytica by activated macrophages depends on the production of reactive nitrogen intermediates which leads to critical iron loss and protozoan parasite death.

Entamoeba histolytica Extract and Interferon-Gamma Activation of Macrophage-Mediated Amoebicidal Function

The effect of recombinant murine interferon-gamma (IFN-gamma) and E. histolytica extract (E.h.E.) on macrophage (M phi) activation for amoebicidal activity was examined. Peritoneal macrophages were harvested from C57BL/6 and A/J mice and preincubated with IFN-gamma and/or E.h.E. It was found that amoebicidal activity could be induced in both C57BL/6 and A/J-derived macrophages by pretreatment with IFN-gamma and E.h.E. Pretreatment of the M phi with E. histolytica extract or IFN-gamma alone did not result in the activation of significant cytotoxic activity against E. histolytica trophozoites. In the presence of IFN-gamma, E.h.E. had a dose-dependent effect on the activation of M phi amoebicidal function.

Passive transfer of immunity against hepatic amoebiasis in the hamster by cells

Summary The role of passive cell‐mediated transfer of immunity in hepatic amoebiasis in hamsters was studied. The transfer of peritoneal cells from hamsters vaccinated against or protected from hepatic amoebiasis and from those with hepatic amoebiasis, as well as of spleen cells from vaccinated or protected, but not from infected hamsters, conferred immunity against hepatic amoebiasis in recipient normal hamsters. Treatment of the spleen cells from protected hamsters with anti T‐cell serum abolished their ability to transfer immunity. It appears that the effector mechanism in this system is T‐cell dependent.

Human monocyte tumouristatic ability: Modulation by cytokines and tumour cell products

The ability of human monocytes to exert cytostatic effects against a murine mastocytoma was assessed. Furthermore, the capacity of different cytokines to endow monocytes with tumouristatic activity was investigated. Overall, our results suggest that human monocytes have a rather high baseline tumouristatic ability against tumours but this varied with experimental conditions, notably the effector to target ratio. This baseline activity was enhanced by treatment with a variety of cytokines, notably interferon-gamma, tumour necrosis factor alpha and granulocyte-macrophage colony stimulating factor. As described in other systems, the best positive results were obtained when combinations of cytokines were used, when synergistic effects were evident. Of interest was the fact that treatment of monocytes with tumour cell products diminished their ability to exert tumouristatic effects against the murine mastocytoma. The importance of these findings to tumour growth in vivo is discussed.

Cellular and Cytokine Profiles in Spontaneous Regression Phase of Hypersensitivity Pneumonitis

The phase of spontaneous regression of hypersensitivity pneumonitis was evaluated using a mouse model. C57BL/6 mice were instilled intranasally with 150 micrograms of the thermophilic actinomycete Faeni rectivirgula 3 days a week so as to establish a mouse model of farmers lungs. It was shown that instillation of mice for a period of more than 6 weeks was associated with a significant decrease in the lung inflammation, suggestive of the so-called spontaneous regression phase seen in this pathology. Indeed, the lung index was seen to decrease after more than 6 weeks of treatment (2.2 after 6 weeks vs. 1.7 at 12 weeks, p < .01). There was also a significant decrease in lung hydroxyproline levels in animals given 12 weeks of treatment (175 micrograms/lung) compared to 6-week-treated animals (212 micrograms/lung, p < .05). Treated mice did not show a significant decrease in the alveolitis after 9 weeks of treatment. Also, there was no evidence that there was a decrease in bronchoalveolar lavage macrophage or T lymphocyte activity in mice given more than 9 weeks of F. rectivirgula treatment, as judged by O2- release and antigen-driven proliferation. Conversely, it was shown that NK cell activity in the lung digest of mice given 9 to 12 weeks of instillation was significantly higher than that seen in mice given 6 weeks of treatment. Analysis of the lung cell cytokine profile seen after ConA mitogenesis showed that after 6 weeks of F. rectivirgula treatments, nonparenchymal cells secreted high levels of tumor necrosis factor alpha (TNF alpha) and granulocyte macrophage colony-stimulating-factor (GM-CSF), whereas similar cells from the lungs of mice given 9-12 weeks of treatment secreted larger amounts of interferon-gamma (IFN gamma) and interleukin-2 (IL-2). Overall, these results suggest that the spontaneous regression phase is associated with changes in NK cell activity and lung cell lymphokine profile.