CXCR4 and MIF are required for neutrophil extracellular trap release triggered by Plasmodium-infected erythrocytes

Neutrophil extracellular traps (NETs) evolved as a unique effector mechanism contributing to resistance against infection that can also promote tissue damage in inflammatory conditions. Malaria infection can trigger NET release, but the mechanisms and consequences of NET formation in this context remain poorly characterized. Here we show that patients suffering from severe malaria had increased amounts of circulating DNA and increased neutrophil elastase (NE) levels in plasma. We used cultured erythrocytes and isolated human neutrophils to show that Plasmodium-infected red blood cells release macrophage migration inhibitory factor (MIF), which in turn caused NET formation by neutrophils in a mechanism dependent on the C-X-C chemokine receptor type 4 (CXCR4). NET production was dependent on histone citrullination by peptidyl arginine deiminase-4 (PAD4) and independent of reactive oxygen species (ROS), myeloperoxidase (MPO) or NE. In vitro, NETs functioned to restrain parasite dissemination in a mechanism dependent on MPO and NE activities. Finally, C57/B6 mice infected with P. berghei ANKA, a well-established model of cerebral malaria, presented high amounts of circulating DNA, while treatment with DNAse increased parasitemia and accelerated mortality, indicating a role for NETs in resistance against Plasmodium infection.
Author summary Protozoans of the Plasmodium genre infect red blood cells and cause malaria in humans and various other mammalian species. Estimated malaria cases are at more than 200 million, with 450,000 deaths per year, being cerebral malaria a serious complication that accounts for the majority of deaths. Neutrophils are cells that participate in host defense against pathogens. These cells use various mechanisms to kill invading microrganisms, including the release of webs of DNA, called neutrophil extracellular traps (NETs). These NETs can help control infections but can also induce tissue damage and their role in malaria and the mechanisms of NET production during malaria infection are starting to be understood. Here we show that infected red blood cells produce a cytokine, macrophage migration inhibitory factor (MIF) that stimulates neutrophils to release NETs. These NETs function to limit Plasmodium dissemination and, thus, digestion of NETs with DNAse treatment causes increased parasitemia and accelerated death in an experimental model of cerebral malaria. Our study uncovers the mechanism by which infected red blood cells stimulate neutrophils to release NETs and suggest an important participation of this process in malaria control.