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Pretreatment with phorbol myristate acetate inhibits macrophage activity against intracellular protozoa
Abstract:
To further document the role of toxic oxygen intermediates in mononuclear phagocyte antiprotozoal activity, microbicidal macrophages were depleted of the capacity to generate superoxide anion (O-2) and hydrogen peroxide (H2O2) by pretreatment with phorbol myristate acetate (PMA), a soluble agent which triggers the macrophage respiratory burst. Treating cells for 90 min with 200 ng/ml of PMA inhibited the extracellular release of both O-2 and H2O2 by 90% upon subsequent restimulation with either PMA or opsonized zymosan. This effect persisted for 48 h, and could not be reversed by the addition of lymphokine. Intracellular nitro-blue tetrazolium reduction by PMA-treated cells was also inhibited by 66-95% upon rechallenge with either PMA or inert or viable particulate agents. In parallel, PMA pretreatment abolished or markedly impaired the ability of normal, lymphokine-stimulated, and in vivo activated macrophages to kill three diverse protozoa--Toxoplasma gondii, Leishmania donovani, and Trypanosoma cruzi. These studies illustrate an additional technique for investigating the antiprotozoal effects of macrophage-derived O-2 and H2O2 and reemphasize the importance of an intact respiratory burst mechanism in killing of intracellular parasites.
Insights
Phorbol myristate acetate (PMA) pretreatment inhibits macrophage production of superoxide anion and hydrogen peroxide. This impairment blocks the killing of protozoa like Toxoplasma gondii, Leishmania donovani, and Trypanosoma cruzi.
Area of Science:
- Immunology
- Cell Biology
- Parasitology
Background:
- Mononuclear phagocytes play a crucial role in combating protozoal infections.
- Toxic oxygen intermediates, such as superoxide anion and hydrogen peroxide, are implicated in the microbicidal activity of macrophages.
Purpose of the Study:
- To investigate the role of superoxide anion (O-2) and hydrogen peroxide (H2O2) in the antiprotozoal activity of macrophages.
- To document the effect of inhibiting the macrophage respiratory burst on protozoal killing.
Main Methods:
- Macrophages were pretreated with phorbol myristate acetate (PMA) to inhibit superoxide anion and hydrogen peroxide generation.
- Inhibition of O-2 and H2O2 release was assessed upon restimulation with PMA or opsonized zymosan.
- Nitro-blue tetrazolium reduction was used to measure intracellular oxidative activity.
- The killing of three protozoan species (Toxoplasma gondii, Leishmania donovani, Trypanosoma cruzi) by treated macrophages was evaluated.
Main Results:
- PMA pretreatment inhibited extracellular O-2 and H2O2 release by 90% for at least 48 hours.
- Intracellular nitro-blue tetrazolium reduction was significantly inhibited (66-95%) in PMA-treated cells.
- PMA pretreatment abolished or markedly impaired the ability of macrophages to kill Toxoplasma gondii, Leishmania donovani, and Trypanosoma cruzi.
Conclusions:
- An intact macrophage respiratory burst mechanism, involving O-2 and H2O2, is essential for killing intracellular parasites.
- PMA-induced inhibition of the respiratory burst provides a method to study the antiprotozoal effects of these oxygen intermediates.
- These findings reemphasize the importance of toxic oxygen intermediates in macrophage-mediated host defense against protozoa.