Blastocystis hominis and the evaluation of efficacy of metronidazole and trimethoprim/sulfamethoxazole.

Blastocystis hominis is commonly found in the intestinal tract of humans. Although the pathogenicity of this unicellular parasite is controversial, anti-protozoan agents are usually administered to infected individuals. At present, the first choice of chemotherapeutic agent is Metronidazole as described in the literature. In this study, we evaluated the effects of metronidazole and Trimethoprim/Sulfamethoxazole (TMP/SMX) on persons infected with B.hominis. A total of 104 subjects infected with B. hominis were admitted to the laboratory from 2002 to 2003. All individuals were non-immunocompromised and subjects were monitored for 1 year after treatment. All stool samples were microscopically examined after staining with iodine and by culturing in an egg slant medium. Of the 104 infected individuals (52+/-16 years of age, M:F=60:44) with B. hominis infection, 28 were discharging large numbers of parasites before treatment. Of 28 severely infected individuals, 12 were treated with metronidazole/250-750 mg at a regimen of 3 x/day/10 days and 4 of the 12 were eradicated. Nine individuals were treated with TMP/SMX/1 tab at a regimen of 3 x/day/10 days and 2 of the 9 were eradicated. For severe B. hominis infections, it appears that metronidazole and TMP/SMX are effective in some individuals, but not all.

DNA-mobility shift assay and the detection of anti-DNA IgG in systemic lupus erythematosus patients.

Although the presence of antibodies against double-stranded (ds) DNA is highly specific of systemic lupus erythematosus (SLE), it is not detected in all SLE patients, perhaps due to a lack of sensitivity of the tests routinely used to assay anti-(ds) DNA. Looking for an alternative assay, this study explored the applicability of a DNA-mobility shift assay for the detection of anti-(ds) DNA; furthermore, the study compared the use of Salmonella typhimurium DNA with that of calf thymus DNA in the assay. After electrophoresis, samples containing S. typhimurium DNA and IgG from SLE sera showed marked alterations in DNA electrophoretic mobility when compared to DNA alone. In our sampling, SLE patients who tested negative for anti-(ds) DNA antibodies with routinely used assays such as Crithidia luciliae immunofluorescence test, radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA), tested positive for anti-(ds) DNA with the DNA mobility shift assay using S. typhimurium DNA. Incubation with IgG from control sera in the same proportions as above did not affect S. typhimurium DNA electrophoretic mobility. When S. typhimurium DNA was replaced by calf thymus DNA, the effect on the DNA mobility was less pronounced and less reliable. These results indicated that a DNA-mobility shift assay would be a useful alternative for the unequivocal detection of abnormal titers of anti-(ds) DNA antibodies. Furthermore, data indicated a greater ability of the IgG from SLE patients to form complexes with S. typhimurium DNA than with calf thymus DNA, suggesting an alternative testing DNA which may lead to a more sensitive anti-(ds) DNA detection.

Dysregulation of interleukin 8, interleukin 10, and interleukin 12 release by alveolar macrophages from HIV type 1-infected subjects.

We examined the in vitro release of interleukin 8 (IL-8), interleukin 10 (IL-10), and interleukin 12 (IL-12) by alveolar macrophages from normal volunteers and HIV-1-infected subjects. Normal volunteers had very low levels of IL-8 and IL-10 and undetectable IL-12 in the cell-free bronchoalveolar lavage fluid (BALF). Asymptomatic HIV-1-infected subjects had elevated levels of IL-8 and IL-10 in their BALF, and HIV-1-infected subjects with nonspecific interstitial pneumonitis (NIP) or infected with Pneumocystis carinii had the highest BALF levels of IL-10 and IL-8. It was found that alveolar macrophages from asymptomatic HIV-1 subjects and from NIP subjects spontaneously released elevated IL-8, IL-10, and IL-12. However, AIDS subjects infected with P. carinii had cells that released elevated levels of IL-10 and IL-8, but low levels of IL-12. When alveolar macrophages were stimulated with Staphylococcus aureus Cowan (SAC), cells from normal volunteers responded with a considerably increased release of IL-8, IL-10, and IL-12; cells from HIV-1-infected subjects without P. carinii infection responded with a moderate increase in release of all three monokines. SAC stimulation did not enhance the release of monokines by cells from AIDS subjects with P. carinii infection, and IL-12 levels remained low. There was no strict relationship between spontaneous cytokine release and p24 HIV-1 antigen expression by alveolar macrophages. Finally, we showed that neutralizing IL-10 production by alveolar macrophages from AIDS subjects substantially increased IL-12 releasability.(ABSTRACT TRUNCATED AT 250 WORDS)

Interaction between Mycobacterium avium and human immunodeficiency virus type 1 (HIV-1) in bronchoalveolar macrophages of normal and HIV-1-infected subjects.

Bronchoalveolar lavage (BAL) macrophages from patients with symptomatic or asymptomatic HIV-1 infections were obtained, and their ability to restrict in vitro the growth of an AIDS-associated strain of Mycobacterium avium was compared with cells obtained from normal volunteers. BAL macrophage populations from HIV-1-infected subjects (symptomatic or asymptomatic) spontaneously released significant amounts of IL-6, IL-1 beta, and TNF-alpha, whereas BAL macrophages from normal volunteers released very low amounts of these cytokines. Phagocytosis of M. avium was shown to be similar in both HIV-1-infected subjects and in control subjects. BAL macrophages from HIV-1-infected subjects released significantly greater quantities of IL-6, IL-1 beta, and TNF-alpha than did cells from normal volunteers upon M. avium ingestion. Growth of M. avium was similar in BAL macrophages from all three subject groups. Finally, BAL macrophages from normal volunteers were obtained, and these cells were doubly infected with a macrophage tropic isolate of HIV-1 at a low multiplicity of infection and with an AIDS-associated strain of M. avium. There were no significant differences in cytokine release by cells co-infected with M. avium and HIV-1 and cells infected with M. avium alone. The growth of mycobacteria and the viral replication in doubly infected cells were compared with those in cells infected with only one of the pathogens, and it was shown that HIV-1 infection had no significant effect on M. avium growth.(ABSTRACT TRUNCATED AT 250 WORDS)

Mycobacterium avium infection in HIV-1-infected subjects increases monokine secretion and is associated with enhanced viral load and diminished immune response to viral antigens.

The complex interaction between HIV-1 infection and Mycobacterium avium was studied. Viral burden was assessed, as well as immune response to HIV-1 in the context of Myco. avium infections. We also examined serum cytokine levels and cytokine release by blood mononuclear cells in HIV-1-infected subjects, infected or not with Myco. avium. Undetectable serum levels of IL-1, tumour necrosis factor-alpha (TNF-alpha) and IL-6 were found in normal controls and in groups I, II and III of HIV-1-infected subjects. Moderate levels of TNF-alpha, IL-1 and IL-6 were found in the sera of group IV patients. When group IV was subdivided into subjects with and without Myco. avium infections, subjects with Myco, avium infections were shown to have higher serum levels of TNF-alpha, IL-1 beta and IL-6 than those with other infections. Blood mononuclear cells from controls and HIV subjects were stimulated with bacterial lipopolysaccharide, and cytokine levels assessed. Cells from group II patients were shown to secrete normal levels of TNF-alpha and IL-6, and lower levels of IL-1 beta; group III subjects released higher levels of IL-6. Patients in group IV had blood cells that released elevated levels of IL-6 and TNF-alpha, and lower levels of IL-1 beta. Group IV subjects with Myco. avium infections had blood cells that released higher levels of TNF-alpha, IL-6 and IL-1 than group IV subjects with other infections. Assessment of viral burden in cells of HIV-1-infected subjects revealed that Myco. avium-infected subjects had a higher level of virus burden and a lower level of lymphoproliferative response to an inactivated gp120-depleted HIV-1 antigen than AIDS subjects with other infections. These data suggest that Myco. avium infections in HIV-1-infected subjects hasten the progression of viral disease, enhance cytokine release and contribute to the anergy to viral antigens.

Interleukin 13 and interleukin 4 protect bronchoalveolar macrophages from productive infection with human immunodeficiency virus type 1.

In this study, we examined the impact of the predominantly Th2-type lymphokines interleukin 13 (IL-13) and interleukin 4 (IL-4) on acute infection of human bronchoalveolar macrophages with a macrophage-tropic isolate of human immunodeficiency virus type 1 (HIV-1). Addition of 0.01-10 ng of IL-4 or IL-13 per milliliters significantly blocked HIV-1 replication in infected cells, judging from levels of reverse transcriptase and p24 antigen in the supernatants of infected cells. Both IL-4 and IL-13 were almost as efficient as interferon-gamma (IFN-gamma) in preventing HIV-1 replication, when given in equivalent amounts. Moreover, neither IL-13 nor IL-4 interfered with the IFN-gamma-mediated enhancement of anti-HIV-1 activity in alveolar macrophages. Both IL-4 and IL-13 interfered with enhanced replication of HIV-1 in macrophages pulsed with the growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF). Interleukin 13 also prevented HIV-1 release from peripheral blood mononuclear cells in a cocultivation experiment with feeder cells from a seronegative subject. These data suggest that Th2-derived lymphokines have significant anti-HIV-1 activity in cells of the macrophage lineage, although they may enhance the susceptibility of HIV-1-infected subjects to some opportunistic pathogens.

Alveolar macrophages from subjects infected with HIV-1 express macrophage inflammatory protein-1 alpha (MIP-1 alpha): contribution to the CD8+ alveolitis.

We examined the synthesis and release of MIP-1 alpha in alveolar macrophages obtained from normal subjects or subjects infected with HIV-1, at different stages of the disease. HIV-1-infected subjects in groups II, III and IV all had significant interstitial pneumonitis, featuring a significant infiltration of CD8+ lymphocytes in the bronchoalveolar lavage. Alveolar macrophages from HIV-1-infected subjects were shown to express significant levels of MIP-1 alpha via immunohistochemistry, both spontaneously and in response to lipopolysaccharide (LPS), whereas cells from normal subjects expressed very low levels of the cytokine. Supernatants of alveolar macrophages from HIV-1-infected subjects exerted strong chemotactic activity for purified activated blood CD8+ T lymphocytes, which was strongly inhibited by neutralizing MIP-1 alpha. Studies of patients with HIV-1 infection at different stages of the disease showed that MIP-1 alpha secretion increased as viral infection developed. There was a significant positive correlation between MIP-1 alpha secretion and the CD8+ alveolitis in HIV-1-infected subjects. Infection of alveolar macrophages in vitro with three distinct strains of HIV-1 which replicated profusely in macrophages did not induce the expression of MIP-1 alpha. Collectively, these data suggest that HIV-1 infection in vivo induces MIP-1 alpha expression and release in alveolar macrophages, and this appears to contribute significantly to the alveolar lymphocytosis seen in HIV-1-infected subjects.

Interleukin-1 is involved in mouse resistance to Mycobacterium avium.

In this study, we examined the contribution of the monokine interleukin-1 (IL-1) in mouse resistance to the intracellular pathogen Mycobacterium avium. The effect of neutralizing endogenous IL-1 in mouse macrophage resistance to M. avium infection was investigated. Infection of mouse peritoneal macrophages with M. avium B101 was shown to result in significant IL-1 beta release by cells at 4 and 7 days postinfection. Addition of IL-1 receptor antagonist (IL-1ra) at doses of 5 micrograms daily, which neutralized endogenous IL-1, failed to significantly modify the intracellular growth of M. avium. Mice were injected with M. avium B101 by the intravenous route, and the growth of the mycobacteria was monitored in the organs of intact mice and in those of mice that received repeated high doses of IL-1ra. The infection with M. avium elicited the production of large amounts of IL-1 in the lungs, livers, and spleens. Repeated injections of IL-1ra into M. avium-infected mice resulted in moderately enhanced growth of the bacilli in the livers and spleens but in much enhanced growth in the lungs. The enhanced growth of M. avium in the lungs correlated with a diminished inflammatory influx of cells (particularly neutrophils) in the bronchoalveolar space. These data argue for a role for IL-1 in host resistance to M. avium infections.

IL-10 neutralization augments mouse resistance to systemic Mycobacterium avium infections.

In this contribution, we examined the involvement of the cytokine IL-10 in the progression of experimental murine Mycobacterium avium infections in susceptible BALB/c mice. Addition of anti-IL-10 antibodies in the supernatants of peritoneal macrophages infected with virulent M. avium resulted in a significantly enhanced mycobacteriostatic activity of macrophages. In BALB/c mice infected with the B101 or B102 virulent M. avium strains, examination of the cytokine release profile in splenocytes from infected mice showed that infection was associated with an initial copious release of both IFN-gamma and IL-10. IL-10 production increased as the infection progressed, whereas IFN-gamma levels diminished. Infected mice were given repeated infusions of a rat mAb against mouse IL-10 or rat IgM. Examination of IgM serum levels in anti-IL-10-treated mice (infected or not) showed that depletion of endogenous IL-10 resulted in much decreased IgM levels. Results showed that infusions of large dosages of the monoclonal anti-IL-10 resulted in a very significantly diminished bacterial growth in the spleens. These findings indicate that IL-10 may have a negative impact on resistance to M. avium infections, due, at least in part, to decreased macrophage activity.

In vitro effect of recombinant interferon gamma in combination with LPS on amoebicidal activity of murine Kupffer cells.

The present study examines the role of liver macrophages (Kupffer cells), of C57BL/6 mice, as effector cells responsible for the killing of Entamoeba histolytica trophozoites in vitro. It was shown that unstimulated Kupffer cells were inefficient in the killing of E. histolytica trophozoites in vitro. Interferon gamma (IFN-gamma) alone was not able to activate Kupffer cells to amoebicidal state. However, Interferon gamma and lipopolysaccharide (LPS) acted synergistically in this phenomenon. It seems that the acquisition of amoebicidal activity is associated with the involvement of hydrogen peroxide, because the addition of catalase partially decreases the killing of this parasite by Kupffer cells. In addition, it appears that the amoebicidal activity of IFN-gamma-treated Kupffer cells is contact-dependent. Our results indicate that the immunologic production of IFN-gamma is important in the activation of Kupffer cells for controlling this parasite and that Kupffer cells are strong effector cells against the amoebae.

Effect of recombinant tumor necrosis factor alpha on in vitro amoebicidal activity of murine Kupffer cells.

The present study was undertaken to determine whether recombinant tumor necrosis factor alpha (TNF) could induce Kupffer cells to kill Entamoeba histolytica parasite in vitro. C57BL/6 mice were used in this study. The liver was perfused and Kupffer cells harvested and treated with TNF for 6 h. It was found that Kupffer cells treated with TNF are able to kill amoebic trophozoites in vitro. These results further show that amoebicidal activity of TNF-activated Kupffer cells is dependent on the ratio of Kupffer cells to amoebic trophozoites. The maximum amoebicidal activity of Kupffer cells was observed with the ratio of one Kupffer cell to five amoebae. This study also shows that the optimal concentration of TNF is required in the induction of amoebicidal activity in Kupffer cells (10(5) units). It seems that both oxidative-dependent and -independent mechanisms are important for the killing of amoebae by the TNF-treated Kupffer cells. It is likely that TNF-treated Kupffer cells produce endogenous TNF or other cytotoxic molecules which are capable of mediating the parasite killing. Our results indicate that the immunologic production of TNF is important in the activation of Kupffer cells to kill amoebic trophozoites.

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 farmer's 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.

Immunotherapy of airborne tuberculosis in mice via the lung-specific delivery of cytokines.

The immunotherapeutic potential of interleukin-2 (IL-2), tumour necrosis factor alpha (TNFα) and interferon gamma (IFN-γ) administered by aerosol was examined on mice infected with Mycobacterium tuberculosis by the aerogenic route. Infection of balb/c mice with 10(4) colony forming units (cfu) of M tuberculosis led to death of all mice at day 35 post infection after progressive microbial growth in the lungs. Aerosolization of IL-2 (100 µg per mouse) did not promote an increase in resistance to tuberculosis, as seen by growth of M tuberculosis in the lungs. Administration of IFN-γ or TNFα (100 µg) by the aerosol route led to a significant reduction in microbial growth in the lungs and a 100% survival of infected mice at day 60. Similarly, aerosolization of TNFα and IFN-γ combined led to a very high degree of tuberculostatic activity in the lungs of infected animals, but not superior to that seen with either cytokine alone. Administration of similar amounts of cytokines by repeated intraperitoneal infusions led to a very marginal improvement in mouse resistance. These results suggest that localized cytokine administration may be beneficial in the treatment of lung diseases.

Murine hypersensitivity pneumonitis: production and importance of colony-stimulating factors in the course of a lung inflammatory reaction.

The release of colony-stimulating factors (CSFs) and their contribution to the inflammatory response in the lungs of mice exposed by the intranasal route to the actinomycete Faeni rectivirgula (150 micrograms/day, 3 days/wk), an important thermophilic actinomycete that determines farmer's lung in humans, was examined. Bronchoalveolar lavages (BAL) and lung homogenates of normal mice or saline-instilled mice contained undetectable levels (less than 0.5 U/ml) of the cytokines interleukin-3 (IL-3), colony-stimulating factor-1 (CSF-1), and granulocyte/macrophage colony-stimulating factor (GM-CSF). Mice instilled with F. rectivirgula developed a CSF cytokine response early (24 h) after the instillation that increased and plateaued 2 wk later, and stayed high thereafter. Similarly, lung homogenates of F. rectivirgula-challenged mice contained significant levels of all three CSFs from 24 h after treatment until termination of the experiment. The offending agent itself, F. rectivirgula, was found to directly induce the secretion of IL-3 and GM-CSF from isolated mouse BAL cells and mouse splenocytes, at doses ranging from 1 to 100 micrograms/ml. This was not due to contaminating endotoxin, as inclusion of polymyxin B did not modify this release. Instillation of antibodies against the CSFs in mice challenged with F. rectivirgula did not modify the increase in BAL cell number determined by the challenge (11-fold increase in BAL cell number in F. rectivirgula-instilled mice at 3 wk, whether given anti-CSFs or not). Moreover, direct intratracheal infusion of CSFs (5,000 U of IL-3/CSF-1/GM-CSF) every week did not change the cellular response seen in challenged mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Murine hypersensitivity pneumonitis: bidirectional role of interferon-gamma.

C57BL/6 mice were instilled intranasally with optimal doses [150 micrograms of antigen 3 days a week) of the actinomycete Faeni rectivirgula to induce an experimental hypersensitivity pneumonitis. Some control mice received normal rat IgG as controls, whereas other mice received 1 mg weekly of rat anti-murine interferon gamma (IFN-gamma) antibody by the intraperitoneal route and 200 micrograms by the intranasal route given 2 days before and during the challenge period before each instillation. Control mice developed a clear hypersensitivity pneumonitis characterized by an early neutrophilic response at 3 days and a later influx of mononuclear cells (nine- to tenfold increase in cell number. P less than 0.001 vs saline instilled mice at 4 weeks post-treatment). F. rectivirgula instillation determined a sharp increase in the lung index (80% increase in lung weight, P less than 0.005 vs saline treated mice), as well as a significant fibrosis at 4 weeks (twofold increase in lung hydroxyproline levels). Cytokine measurements showed that tumour necrosis factor alpha (TNF alpha) was present in the broncho-alveolar lavage (BAL) of challenged mice at 4 weeks when the BAL was obtained 8 hr after the last challenge (130 U/ml). Treatment of mice with the monoclonal antibody against IFN-gamma was associated with very few changes in the number of cells in the BAL of challenged mice. The lung index of challenged mice was significantly reduced by infusion of the anti-IFN-gamma antibody. Anti-IFN-gamma treatment resulted in decreased levels of TNF alpha in the BAL of F. rectivirgula after 4 weeks of treatment (56 U/ml, P less than 0.01). Moreover, depletion of endogenous IFN-gamma in F. rectivirgula-instilled mice resulted in a diminished lung fibrotic response (P less than 0.01 vs mice treated with F. rectivirgula and control antibody). We also studied the effect of exogenous IFN-gamma adminstration on the development of lung disease. Groups of mice received recombinant gamma interferon (IFN-gamma) (1000 U) intraperitoneally just before the first treatment and also daily, whereas controls received saline or IFN-gamma alone (no F. rectivirgula challenge). After 4 weeks of treatment, mice were killed and various markers of the disease were evaluated. As mentioned before, bronchoalveolar lavage (BAL) cell number was increased tenfold in mice treated with F. rectivirgula, whereas mice given F. rectivirgula and IFN-gamma had only a threefold increase in BAL cell number, determined mostly by a decrease in alveolar macrophage recruitment in the lungs.(ABSTRACT TRUNCATED AT 400 WORDS)

Transforming growth factor-beta is generated in the course of hypersensitivity pneumonitis: contribution to collagen synthesis.

Mice of the C57BL/6 strain were instilled with optimal doses (150 micrograms/day for 3 days/wk) of the thermophilic actinomycete Faeni rectivirgula (also known as Saccharopolyspora rectivirgula or Micropolyspora faeni) to induce a hypersensitivity pneumonitis inflammation that mimics the human disease affecting certain occupational groups. This mouse model was characterized by a very significant alveolitis (3-fold increase in bronchoalveolar lavage [BAL] cell number at 48 h and a 10-fold increase at 3 wk). Also, total lung transforming growth factor (TGF-beta) was shown to be elevated in treated mice as early as 1 wk after the first instillation and increased gradually to 2.5 micrograms/lung at 3 wk (approximately 0.3 microgram/lung in saline-instilled controls). Intranasal instillation with F. rectivirgula was also associated with very significant increases in lung fibroblast collagen synthesis, starting at 2 wk. BAL macrophages from mice instilled with F. rectivirgula were found to release significantly more TGF-beta upon in vitro stimulation with zymosan beads than did BAL macrophages from saline-instilled mice. Finally, we show that supernatants from activated BAL macrophages of mice given F. rectivirgula increased quite significantly collagen synthesis in normal mouse lung fibroblasts. This increase could be abrogated by treating conditioned medium with a rabbit antibody against TGF-beta. Collectively, these data suggest that TGF-beta is generated in the course of experimental mouse hypersensitivity pneumonitis and contributes significantly to collagen synthesis.

In vivo activation of macrophages by IFN-gamma to kill Entamoeba histolytica trophozoites in vitro.

To determine the role of interferon-gamma (IFN-gamma) in the activation of macrophages to kill Entamoeba histolytica trophozoites in vitro, C57BL/6 mice were injected with various doses of recombinant IFN-gamma (rIFN-gamma) by either the intravenous, intraperitoneal or intramuscular routes. Mice were treated with doses of rIFN-gamma ranging from 10(1) to 10(5) 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-gamma, 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-gamma 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-gamma can activate murine peritoneal macrophages to kill amoebic trophozoites.

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.

Murine hypersensitivity pneumonitis: interleukin-4 administration partially abrogates the disease process.

C57BL-6 inbred mice were given intranasal instillations of Faeni rectivirgula (150 micrograms/day, 3 days a week for 3 weeks) to produce a lung inflammatory reaction which mimics Farmers' lung in humans. Challenged mice developed a strong inflammatory response in their lungs, based on various markers (lung index, bronchoalveolar cell number, fibrosis). The effect of interleukin-4 (IL-4) was studied by infusing mice intraperitoneally with 100, 1000 or 10,000 units of IL-4 weekly during the challenge period. It was shown that IL-4 infusion decreased the inflammatory response, as seen by a decreased lung index (1.7 in mice given F. rectivirgula and 10(3) U IL-4 and 1.31 in mice given antigen and 10(4) U IL-4 weekly versus 2.3 in mice instilled with F. rectivirgula). Interleukin-4 infusion also partially abrogated the F. rectivirgula-induced alveolitis, as seen by a decrease in cell numbers in the broncho-alveolar lavage (BAL) (1.3 x 10(5) cells in saline-instilled mice; 8.3 x 10(5) cells in mice given 10(3) U IL-4 and F. rectivirgula; 3.2 x 10(5) cells in mice given antigen and 10(4) U IL-4; 1.8 x 10(6) cells in mice given F. rectivirgula only). Also, it was apparent that IL-4 administration could partially block the appearance of the fibrosis induced by F. rectivirgula (220 micrograms of hydroxyproline/lung in challenged mice; 170 micrograms/lung in challenged mice given 10(3) U IL-4; 131 micrograms/lung in mice given antigen and 10(4) U IL-4 and c. 100 micrograms/lung in control animals). Infusion of 10(2) U IL-4 weekly had no statistical effect on any marker of inflammation.(ABSTRACT TRUNCATED AT 250 WORDS)

Activated mouse macrophages kill Entamoeba histolytica 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.