Effect of Imiquimod on Chronic Toxoplasmosis and its Associated Neurological Complications in a Rat Model

Abstract

Background: Toxoplasma gondii (T. gondii) is the etiologic agent of toxoplasmosis, a prevalent protozoan parasitosis infecting one-third of the human population worldwide and more than two-thirds of the Lebanese population. T. gondii targets primarily the brain in which, under the host immune control, tachyzoites responsible for acute toxoplasmosis (AT), transform into latent bradyzoites responsible for chronic toxoplasmosis (CT). In immunocompromised patients, CT may reactivate leading to a potentially life-threatening condition. Recently, several associations between CT and behavioral neurological complications were highlighted. However, a direct pre-clinical or clinical implication of toxoplasmosis with most of these diseases remains elusive. Furthermore, an approved treatment targeting CT is still lacking despite its high prevalence.

Aims: Our group previously demonstrated the high potency of an immunomodulatory drug, Imiquimod, against AT and CT, and unraveled its molecular mechanism of action against CT. Briefly, Imiquimod induced interconversion from bradyzoites to tachyzoites, upregulated Toll-like receptors, leading to the consequent activation of the MyD88 pathway, triggering the host immune response and the control of reactivated Toxoplasma foci. The aim of the current work is to study the molecular effects of Imiquimod on CT-associated behavioral complications, namely anxiety-like behavior and hippocampal cognitive functions, in a chronically infect rat model with T.gondii. Methods: One-month-old male Sprague Dawley rats were intraperitoneally injected with 3 million 76K strain of T. gondii. One-week post-infection, blood samples were collected and acute toxoplasmosis was verified using immunofluorescence assay and western blot. Seropositive rats were then treated intraperitoneally with Imiquimod, at a dose of 2.5 mg/kg/day every other day for 2 weeks. Rats were divided into 4 groups: a control untreated group (CU), a control group treated with Imiquimod (CT), a group of untreated rats infected with T.gondii (TU), and a group of rats infected with T.gondii treated with Imiquimod (TT). A panel of three behavioral tests (open-field test, forced-swim test, and Morris water maze test) was performed. Finally, rats were sacrificed and their brains were harvested. The number of cysts in the brain of each rat was counted, and transcript levels of different pro-inflammatory and anti-inflammatory cytokines were analyzed by quantitative real-time PCR. Results: We successfully reproduced a rat model with CT. Consistent with our previous results obtained in mouse models, Imiquimod decreased the number of cysts in the brain of chronically infected rats and induced the reactivation of bradyzoites into tachyzoites. Moreover, an upregulation of proinflammatory cytokines as well as of the inducible Nitric Oxide Synthase (i-NOS) in the brains of chronically infected rats was observed upon treatment with Imiquimod, and triggered a concurrent anti-inflammatory immune response to lessen the immune-pathological response. Importantly, in chronically infected rats with T. gondii, Imiquimod treatment alleviated the anxiety-like behavior and reversed Toxoplasma-induced learning deficits in infected rats.

Conclusion: Our results enhance our knowledge on the implications of chronic toxoplasmosis on behavioral aberrancies, and highlight the potency of Imiquimod on these CT-associated neurological complications.