Chronic infection with Toxoplasma gondii can reactivate and cause life-threatening toxoplasmic encephalitis in AIDS patients and other immunocompromised individuals. The basis of persistence of this chronic infection is the tissue cyst. Thus, eliminating T. gondii cysts from chronically infected individuals before they become severely immunosuppressed will be able to prevent this serious disease. However, there are currently no drugs effective against the cysts. Our recent studies using a murine model uncovered that CD8+ immune T cells have a potent activity to directly penetrate into the cysts and induce their elimination through perforin- mediated mechanisms. We also identified that the N-terminal region (amino acids 41-152) of dense granule protein (GRA) 6 (GRA6Nt) of the parasite presented by the H-2Ld molecule is a key antigen to activate the anti- cyst CD8+ T cells. Our studies also identified that Iba1+ microglia and Ly6C+ macrophages accumulate to the cysts during the T cell-initiated immune process, and that these phagocytes kill T. gondii bradyzoites within the cysts, at least in part, by phagolysosome acidification. Based on these observations, the proposed studies are designed to obtain the information required for proceeding towards development of a vaccine to activate CD8+ T cells capable of targeting T. gondii cysts and eradicate chronic infection with this parasite in humans. For this purpose, we need to determine whether GRA6Nt activates anti-cyst CD8+ T cells through human MHC class I molecules, since the MHC class I molecules present antigens to activate CD8+ T cells.
Aim 1 will address this point by employing three transgenic mouse strains expressing either of the three major MHC class I supermotifs that cover 90% of human population. Once the anti-cyst T cells are primed, these T cells need to be recruited to cyst-containing cells and activated not only by recognizing T. gondii antigens but also by appropriate costimulatory molecules expressed on the cyst-containing cells. Our recent studies suggested an involvement of CXCL10 chemokine and inducible constimulator ligand (ICOSL) in these processes. Thus, Aim 2 will determine the roles of CXCL10 and ICOSL in recruitment and activation of anti-cyst CD8+ T cells, respectively. Another critical but unsolved issue is that small numbers of cysts persist in the brains of infected mice even in the presence of anti-cyst CD8+ T cells. Our recent study revealed that these persisting cysts express significantly greater levels of mRNA for selected GRA and rhoptry (ROP) proteins when compared to the overall cyst population that exists in the absence of the T cells. Since GRAs and ROPs are secreted from the parasite into infected cells, upregulated secretion of these selected GRAs and ROPs could play crucial roles in manipulating the functions of cyst-containing cells to prevent the attack by CD8+ T cells. Thus, Aim 3 focuses to determine whether these GRAs and ROPs play key roles in the evasion of the cysts from the T cell attack. We will also address the molecular mechanisms underlying this evasion process. The information from these studies will establish the basis for developing a novel method to eradicate chronic T. gondii infection.

Public Health Relevance

Toxoplasma gondii establishes a chronic infection by forming tissue cysts preferentially in the brain, and reactivation of this chronic infection causes life-threatening toxoplasmic encephalitis in immunocompromised individuals such as those with AIDS. The proposed studies aim to identify T. gondii antigen applicable to a vaccine for activating the protective immunity capable of eliminating the cysts in humans and to uncover the mechanisms that T. gondii employs to escape from the protective immunity. The combination of the vaccine and a treatment to block the escape mechanism of the pathogen will allow us to develop a novel method to eradicate this widespread chronic infection in humans.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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HIV Coinfections and HIV Associated Cancers Study Section (HCAC)
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Pesce, John T
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University of Kentucky
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Sa, Qila; Tiwari, Ashish; Ochiai, Eri et al. (2018) Inducible nitric oxide synthase in innate immune cells is important for restricting cyst formation of Toxoplasma gondii in the brain but not required for the protective immune process to remove the cysts. Microbes Infect 20:261-266
Sa, Qila; Ochiai, Eri; Tiwari, Ashish et al. (2017) Determination of a Key Antigen for Immunological Intervention To Target the Latent Stage of Toxoplasma gondii. J Immunol 198:4425-4434
Ochiai, Eri; Sa, Qila; Perkins, Sara et al. (2016) CD8(+) T cells remove cysts of Toxoplasma gondii from the brain mostly by recognizing epitopes commonly expressed by or cross-reactive between type II and type III strains of the parasite. Microbes Infect 18:517-22
Sa, Qila; Ochiai, Eri; Tiwari, Ashish et al. (2015) Cutting Edge: IFN-? Produced by Brain-Resident Cells Is Crucial To Control Cerebral Infection with Toxoplasma gondii. J Immunol 195:796-800
Ochiai, Eri; Sa, Qila; Brogli, Morgan et al. (2015) CXCL9 is important for recruiting immune T cells into the brain and inducing an accumulation of the T cells to the areas of tachyzoite proliferation to prevent reactivation of chronic cerebral infection with Toxoplasma gondii. Am J Pathol 185:314-24
Sa, Qila; Ochiai, Eri; Sengoku, Tomoko et al. (2014) VCAM-1/?4?1 integrin interaction is crucial for prompt recruitment of immune T cells into the brain during the early stage of reactivation of chronic infection with Toxoplasma gondii to prevent toxoplasmic encephalitis. Infect Immun 82:2826-39
Sa, Qila; Woodward, Jerold; Suzuki, Yasuhiro (2013) IL-2 produced by CD8+ immune T cells can augment their IFN-? production independently from their proliferation in the secondary response to an intracellular pathogen. J Immunol 190:2199-207
Sullivan, Adam M; Zhao, Xiaopeng; Suzuki, Yasuhiro et al. (2013) Evidence for finely-regulated asynchronous growth of Toxoplasma gondii cysts based on data-driven model selection. PLoS Comput Biol 9:e1003283
Hester, James; Mullins, Jeremi; Sa, Qila et al. (2012) Toxoplasma gondii antigens recognized by IgG antibodies differ between mice with and without active proliferation of tachyzoites in the brain during the chronic stage of infection. Infect Immun 80:3611-20
Suzuki, Yasuhiro; Sa, Qila; Gehman, Marie et al. (2011) Interferon-gamma- and perforin-mediated immune responses for resistance against Toxoplasma gondii in the brain. Expert Rev Mol Med 13:e31