O papel da NINJ1 na resposta inflamatória mediada por piroptose em Toxoplasma gondii e Neospora caninum

Abstract

The innate immune system is crucial for defending against intracellular pathogens, such as Apicomplexan parasites, including Toxoplasma gondii and Neospora caninum. Programmed cell death (PCD) pathways, like pyroptosis, play a key role in limiting parasite replication and promoting inflammatory responses. Pyroptosis is characterized by inflammasome activation and gasdermin pore formation, leading to plasma membrane permeabilization. Recently, Nerve Injury-Induced Protein 1 (NINJ1) has been identified as a key mediator of terminal membrane rupture during inflammatory cell death. However, its role during Apicomplexan infections remains poorly understood. To address this, the study first conducted a review of PCD pathways in Apicomplexan parasites to identify existing knowledge gaps. It then investigated the role of NINJ1 in T. gondii and N. caninum infections. NINJ1-deficient DC2.4 cells (ΔNinj1) were generated using CRISPR-Cas9, and functional assays were performed to assess membrane permeabilization, cell lysis (via PI uptake and LDH release), and caspase-1/11 activation. Additionally, bone marrow-derived macrophages (BMDMs) from wild-type and caspase-1/11-deficient mice were used to assess inflammasome-dependent membrane permeability. Intracellular parasite burden was analyzed in DC2.4 cells and RAW 2.6.4.2 macrophages, comparing parasite replication in normal and NINJ1-deficient cells. The results showed that NINJ1 expression increases during infection and that its absence enhances parasite replication while significantly reducing cell lysis. Loss of NINJ1 did not affect caspase-1/11 activation or initial membrane permeabilization, but LDH release was greatly reduced, indicating that NINJ1 is essential for the terminal phase of plasma membrane rupture. These findings position NINJ1 as a critical effector linking inflammasome activation to terminal cellular disintegration during Apicomplexan infections.