Toxoplasma gondii is dependent on glutamine and alters migratory profile of infected host bone marrow derived immune cells through SNAT2 and CXCR4 pathways.

The obligate intracellular parasite, Toxoplasma gondii, disseminates through its host inside infected immune cells. We hypothesize that parasite nutrient requirements lead to manipulation of migratory properties of the immune cell. We demonstrate that 1) T. gondii relies on glutamine for optimal infection, replication and viability, and 2) T. gondii-infected bone marrow-derived dendritic cells (DCs) display both "hypermotility" and "enhanced migration" to an elevated glutamine gradient in vitro. We show that glutamine uptake by the sodium-dependent neutral amino acid transporter 2 (SNAT2) is required for this enhanced migration. SNAT2 transport of glutamine is also a significant factor in the induction of migration by the small cytokine stromal cell-derived factor-1 (SDF-1) in uninfected DCs. Blocking both SNAT2 and C-X-C chemokine receptor 4 (CXCR4; the unique receptor for SDF-1) blocks hypermotility and the enhanced migration in T. gondii-infected DCs. Changes in host cell protein expression following T. gondii infection may explain the altered migratory phenotype; we observed an increase of CD80 and unchanged protein level of CXCR4 in both T. gondii-infected and lipopolysaccharide (LPS)-stimulated DCs. However, unlike activated DCs, SNAT2 expression in the cytosol of infected cells was also unchanged. Thus, our results suggest an important role of glutamine transport via SNAT2 in immune cell migration and a possible interaction between SNAT2 and CXCR4, by which T. gondii manipulates host cell motility.

Patterns of Toxoplasma gondii cyst distribution in the forebrain associate with individual variation in predator odor avoidance and anxiety-related behavior in male Long-Evans rats.

Toxoplasma gondii (T. gondii) is one of the world's most successful brain parasites. T. gondii engages in parasite manipulation of host behavior and infection has been epidemiologically linked to numerous psychiatric disorders. Mechanisms by which T. gondii alters host behavior are not well understood, but neuroanatomical cyst presence and the localized host immune response to cysts are potential candidates. The aim of these studies was to test the hypothesis that T. gondii manipulation of specific host behaviors is dependent on neuroanatomical location of cysts in a time-dependent function post-infection. We examined neuroanatomical cyst distribution (53 forebrain regions) in infected rats after predator odor aversion behavior and anxiety-related behavior in the elevated plus maze and open field arena, across a 6-week time course. In addition, we examined evidence for microglial response to the parasite across the time course. Our findings demonstrate that while cysts are randomly distributed throughout the forebrain, individual variation in cyst localization, beginning 3 weeks post-infection, can explain individual variation in the effects of T. gondii on behavior. Additionally, not all infected rats develop cysts in the forebrain, and attenuation of predator odor aversion and changes in anxiety-related behavior are linked with cyst presence in specific forebrain areas. Finally, the immune response to cysts is striking. These data provide the foundation for testing hypotheses about proximate mechanisms by which T. gondii alters behavior in specific brain regions, including consequences of establishment of a homeostasis between T. gondii and the host immune response.

An insult-inducible vector system activated by hypoxia and oxidative stress for neuronal gene therapy.

Gene therapy has demonstrated the protective potential of a variety of genes against stroke. However, conventional gene therapy vectors are limited due to the inability to temporally control their expression, which can sometimes lead to deleterious side effects. Thus, an inducible vector that can be temporally controlled and activated by the insult itself would be advantageous. Using hypoxia responsive elements (HRE) and antioxidant responsive elements (ARE), we have constructed an insult-inducible vector activated by hypoxia and reactive oxygen species (ROS). In COS7 cells, the inducible ARE-HRE-luciferase vectors are highly activated by oxygen deprivation, hydrogen peroxide treatment, and the ROS-induced transcription factor NF-E2-related factor 2 (Nrf2). Using a defective herpes virus, the neuroprotective potential of this inducible vector was tested by over-expressing the transcription factor Nrf2. In primary cortical cultures, expression of the inducible ARE-HRE-Nrf2 protects against oxygen glucose deprivation, similar to that afforded by the constitutively expressed Nrf2. This ARE+HRE vector system is advantageous in that it allows the expression of a transgene to be activated not only during hypoxia but also maintained after reperfusion, thus prolonging the transgene expression during an ischemic insult. This insult-inducible vector system will be a valuable gene therapy tool for activating therapeutic/protective genes in cerebrovascular diseases.