γδ T Cells Kill Plasmodium falciparum in a Granzyme- and Granulysin-Dependent Mechanism during the Late Blood Stage.

Plasmodium spp., the causative agent of malaria, have a complex life cycle. The exponential growth of the parasites during the blood stage is responsible for almost all malaria-associated morbidity and mortality. Therefore, tight immune control of the intraerythrocytic replication of the parasite is essential to prevent clinical malaria. Despite evidence that the particular lymphocyte subset of γδ T cells contributes to protective immunity during the blood stage in naive hosts, their precise inhibitory mechanisms remain unclear. Using human PBMCs, we confirmed in this study that γδ T cells specifically and massively expanded upon activation with Plasmodium falciparum culture supernatant. We also demonstrate that these activated cells gain cytolytic potential by upregulating cytotoxic effector proteins and IFN-γ. The killer cells bound to infected RBCs and killed intracellular P. falciparum via the transfer of the granzymes, which was mediated by granulysin in a stage-specific manner. Several vital plasmodial proteins were efficiently destroyed by granzyme B, suggesting proteolytic degradation of these proteins as essential in the lymphocyte-mediated death pathway. Overall, these data establish a granzyme- and granulysin-mediated innate immune mechanism exerted by γδ T cells to kill late-stage blood-residing P. falciparum.

Platelet Endothelial Cell Adhesion Molecule 1 (CD31) Is Essential for Clostridium perfringens Beta-Toxin Mediated Cytotoxicity in Human Endothelial and Monocytic Cells.

Beta toxin (CPB) is a small hemolysin beta pore-forming toxin (ß-PFT) produced by Clostridium perfringens type C. It plays a central role in the pathogenesis of necro-hemorrhagic enteritis in young animals and humans via targeting intestinal endothelial cells. We recently identified the membrane protein CD31 (PECAM-1) as the receptor for CPB on mouse endothelial cells. We now assess the role of CD31 in CPB cytotoxicity against human endothelial and monocytic cells using a CRISPR/Cas9 gene knockout and an antibody blocking approach. CD31 knockout human endothelial and monocytic cells were resistant to CPB and CPB oligomers only formed in CD31-expressing cells. CD31 knockout endothelial and monocytic cells could be selectively enriched out of a polyclonal cell population by exposing them to CPB. Moreover, antibody mediated blocking of the extracellular Ig6 domain of CD31 abolished CPB cytotoxicity and oligomer formation in endothelial and monocytic cells. In conclusion, this study confirms the role of CD31 as a receptor of CPB on human endothelial and monocytic cells. Specific interaction with the CD31 molecule can thus explain the cell type specificity of CPB observed in vitro and corresponds to in vivo observations in naturally diseased animals.