Identifying human neuron IFN-γ dependent anti-Toxoplasma gondii responses

项目总结:

Project Summary: Toxoplasma gondii is an intracellular parasite that latently infects many hosts, including humans. Successful latent infection requires T. gondii to evade cytokine-induced, cell intrinsic responses and switch from a fast-growing form to a slow-growing encysted form. In humans, this life-long infection occurs in the brain where T. gondii can reactivate in the setting of acquired immune deficiencies. In AIDS patients, toxoplasmic encephalitis is the most common cause of focal brain lesions and can cause prolonged neurologic deficits even after appropriate treatment. Recent studies also suggest that, even in HIV+ patients on effective antiretroviral therapy, persistent T. gondii infection may adversely affect cognition and global immune responses. Despite its clinical importance, we lack a mechanistic understanding of what is actually responsible for T. gondii’s pathogenesis in the CNS, including what enables long-term latent infection. Such understanding is crucial to eventually preventing symptomatic disease in HIV/AIDs patients. Within the CNS, cysts are primarily found in neurons. Based on limited in vitro and in vivo studies, it has been presumed that while infected astrocytes cleared intracellular parasites, neurons did not and thus were the de facto host cell for persistent infection. In the last decade, our pioneering work has questioned this model, including showing that IFN-γ-stimulated neurons mount anti-parasitic defenses. As most of this work has occurred in murine models and neurons, it will have missed human neuron-specific pathways (e.g., humans lack the terminal IFN-γ dependent proteins used by murine cells to kill intracellular parasites). The goal of this grant is to address this gap by establishing robust stem cell derived human neuron models (hPSC neurons) which can be used to mechanistically define human neuron-T. gondii interactions that enable immune evasion and persistence. To accomplish this goal, our will leverage our synergistic expertise in CNS toxoplasmosis (Dr. Koshy) and stem cell biology (Dr. Churko) to generate Cre reporter hPSC neurons and CRISPRi-expressing hPSC neurons (Aim 1) and to begin to define the mechanisms by which cytokine stimulated hPSC-neurons control T. gondii (Aim 2). With the completion of these aims, we will have established and validated tools that will form the essential foundation of a long-term program to molecularly define the neuron-T. gondii interactions that enable control and persistence of T. gondii in human neurons. The work proposed here represents an important first step toward developing human neuron-specific therapies for acute and chronic toxoplasmosis. Such therapies will be of great benefit for the HIV/AIDS population at risk for toxoplasmic encephalitis.