Heme and hemozoin induce platelet cell death through UPR-induced apoptosis and ferroptosis in vivax malaria

Malaria is a highly prevalent infectious disease caused by Plasmodium parasites. Plasmodium intraerythrocytic replication leads to hemolysis-driven intermittent febrile crises in patients. In addition, the lysis of unparasitized red blood cells contributes to anemia and endotoxemia. Because thrombocytopenia is an important feature of vivax and severe falciparum malaria, we hypothesized that increased hemolysis in malaria contributes to severe thrombocytopenia by releasing endogenous and parasite toxins (ie, heme and hemozoin) capable of inducing programmed cell death in platelets. Using complementary biochemical, ultrastructural, pharmacological, and molecular approaches, we examined response to stress and cell death pathways that were elevated in the transcriptome of platelets during vivax malaria and evaluated markers of hemolysis that correlated with thrombocytopenia. We found that heme in plasma from thrombocytopenic vivax malaria, but not nonthrombocytopenic vivax or falciparum malaria, induced platelet cell death ex vivo. Platelet stimulation with heme and hemozoin induced apoptotic and necrotic cell death features, with stronger necrosis triggered by hemozoin. Heme and hemozoin activated apoptotic caspases, but only heme induced calpain-dependent BcL-xL degradation, which was not required for platelet apoptosis. We unmasked a caspase-independent intrinsic apoptosis program mechanism depending on the endoplasmic reticulum stress sensor and unfolded protein response trigger IRE1α. Regarding necrosis, we observed inflammasome activation but not pyroptosis. Instead, we distinguished a necrotic cell death feature consistent with ferroptosis, dependent on lipid peroxidation and regulated by DGAT1/2 enzymes, which was the main pathway for hemozoin-induced thrombocytopenia in vitro. Our results identify novel pathways of regulated cell death in platelets that were associated with thrombocytopenia in malaria and may have potential implications for other hemolytic disorders.