Cerebral malaria (CM), a severe complication of Plasmodium falciparum (Pf) infection, is characterised by pleomorphic endothelial cell (EC) activation, adhesion of infected erythrocytes (IE) leucocytes and platelets to vessel walls and subsequent blockade of post-capillary venules, notably in the brain. We showed that IE and host cells activate brain EC and trigger immunopathological changes, notably via extracellular vesicles (EV).
Here, we characterised different classes of EV released in co-cultures of human brain microvascular EC and Pf-IE, using nanoparticle tracking analysis (NTA), flow cytometry (FC) and vibrational spectroscopy. In this in vitro model of CM, we compared two Pf strains: 3D7, which binds to CD36, and E8B, which binds to ICAM-1. While both parasites dramatically increased annexin V+ microvesicles (MV) release, when compared to normal erythrocytes, no difference was observed between the ICAM-1 binder versus the CD36 binder Pf strain. NTA demonstrated that EC-IE supernates had more vesicles than other groups and were the only condition with EV around 500 nm. Interestingly, NTA revealed that the majority of EV released upon contact between IE and brain EC were in the 100-200 nm range, with E8B parasites inducing significantly more EV than 3D7. Thus, small size EV may be the major player. Vibrational spectroscopy allowed us to identify significant differences in the biomolecular content of EV derived from brain EC co-cultured with the ICAM-1 binder versus the CD36 binder Pf strain. These results provide pivotal insight in the mechanisms of EV formation, as well as in their role in the pathophysiology of CM.