The role of chemokines in T cell migration during Toxoplasmic encephalitis

University Of Pennsylvania, Philadelphia PA

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Abstract

DESCRIPTION (provided by applicant): Toxoplasma gondii causes encephalitis in patients with acquired T cell deficiencies, as protective immunity in the central nervous system (CNS) is dependent on T cells and IFN-gamma. Preliminary data shows that antigen-specific CD8+ T cells in the brain during Toxoplasmic encephalitis exhibit a variety of migratory behaviors including association with an infection-induced network. Based on these findings and additional data, we hypothesize that the IFN-gamma-induced chemokine, CXCL10, and its receptor, CXCR3, are necessary for effective T cell responses to T. gondii and influence T cell migration in the CNS. This hypothesis will be tested in two specific aims. Aim A will focus on the role of IFN-gamma and CXCL10 in facilitating a protective T cell response. In order to determine whether T cells expressing CXCR3 have effector capabilities, fluorescence activated cell sorting and flow cytometry will be utilized to determine which cytokines the T cells produce, whether the CD4+ cells are Th1 or Treg cells, and if CD8+ T cells express cytotoxic molecules. In addition, IFN-y and CXCLIO-blocking antibodies will be used to determine how these molecules affect chemokine expression, T cell number and localization, and anti-parasitic responses in the brain. The effects on CNS T cell populations will be evaluated by flow cytometry, T cell localization will be assessed by immunohistochemistry, and chemokine expression and parasite number will be measured by real-time PCR. In Aim B, the effect of CXCR3 and CXCL10 on T cell migratory behavior will be determined using multi-photon (MP) microscopy. Several parameters will be assessed, including, T cell velocity, directed migration, and contact with the fibrous network. The migratory behavior of GFP-expressing CXCR3-deficient OT-I cells will be compared to mCherry-expressing wildtype OT-I cells using MP microscopy on explanted brain. To assess the role of CXCL10, the migratory behavior of GFP+ antigen-specific CD8+ T cells will be observed following anti-CXCLI 0 treatment and compared to mice receiving control antibodies. Overall, this study aims to identify an additional role for IFN-gamma in mediating chemokine-dependent T cell migration and protective immunity in the central nervous system. PUBLIC HEALTH RELEVANCE: Weak immune systems are unable to prevent the parasite, Toxoplasma gondii, from causing a severe brain infection. The aim of this application is to understand how molecules direct cells of the immune system in the brain in order to destroy the parasite. Findings from these studies may lead to new therapies that will enhance the destruction of parasites.