Phagocytic uptake of oxidized heme polymer is highly cytotoxic to macrophages

Area of Science:

• Immunology
• Pathology
• Toxicology

Background:

• Apoptosis in macrophages contributes to immune suppression and pathology in malaria.
• Phagocytosis of Plasmodium-infected red blood cells (PRBC) induces macrophage apoptosis via cytosolic factors.
• Heme compounds, such as heme polymer and beta-hematin, induce dose-dependent macrophage death.

Purpose of the Study:

• To investigate the synergistic toxic effects of methemoglobin with heme compounds on macrophages.
• To elucidate the mechanisms underlying methemoglobin-mediated potentiation of heme toxicity.
• To explore the potential of targeting methemoglobin peroxidase activity for therapeutic intervention in malaria.

Main Methods:

• Macrophage cell death assays (LC50 determination) with heme compounds and methemoglobin.
• Microscopic analysis (light and scanning electron microscopy) to observe beta-hematin accumulation.
• Assessment of reactive oxygen species (ROS) generation and DNA fragmentation.
• Inhibition studies using spin-trap Phenyl-N-t-butyl-nitrone (PBN).

Main Results:

• Methemoglobin significantly amplified the toxicity of heme polymer and beta-hematin towards macrophages.
• Beta-hematin uptake via phagocytosis was crucial for enhanced toxicity in the presence of methemoglobin.
• Macrophage death was mediated by an oxidative stress-dependent apoptotic pathway, involving H2O2 generation.
• Methemoglobin oxidized beta-hematin to an oxidized form (betaH*) through a single electron transfer mechanism, which was implicated in toxicity.

Conclusions:

• Methemoglobin and pro-oxidant heme molecules cooperatively enhance toxicity towards macrophages.
• The potentiation of toxicity is linked to the generation of oxidized beta-hematin (betaH*) and oxidative stress.
• Targeting methemoglobin peroxidase activity could be a strategy to mitigate the cellular toxicity of pro-oxidant molecules and bolster host immune response against malaria.