Investigating the role of the innate immune system in entrapping and neutralizing Toxoplasma gondii by neutrophil extracellular traps (NETs) 4349

Abstract Description Approximately one-third of the global human population harbors an infection with the protozoan parasite Toxoplasma gondii. The immune system relies on neutrophil extracellular traps (NETs), to entrap and neutralize T. gondii. NETs are comprised of DNA, myeloperoxidase, citrullinated-histone 3, and elastase. Recent findings have suggested an alternative outcome - vital NETosis, which allows NET formation while preserving cell viability. While T. gondii induces NETs, whether these are released through vital NETosis remains. Additionally, the cellular interactions that trigger vital NETosis are unknown. A promyelocytic cell line HL-60 was differentiated into neutrophil-like cells (NLCs) using retinoic acid and DMSO. NLCs or primary human neutrophils were then infected with T. gondii at a multiplicity of infection of 2 in the presence of SYTOX, a dye that binds to extracellular DNA. Fluorescence was quantified with a plate reader and compared with PMA-stimulated cells, an inducer of suicidal NETosis. Results showcased increased extracellular DNA in T. gondii-infected NLCs and primary human neutrophils. Immunofluorescence microscopy at various time points confirmed NET formation. The viability of T. gondii-infected NLC at later time points was confirmed by zombie staining. The next stages of this project are determining whether neutrophil complement receptors mediate T. gondii-induced vital NETosis and the role of RAB27a is associated with vital NETosis. Funding Sources NIH NIGMS T32GM137812, NIH NIAID 1R16AI184345-01 Topic Categories Innate Immune Responses and Host Defense: Cellular Mechanisms (INC)