CXCR3 chemokine receptor guides Trypanosoma cruzi-specific T-cells triggered by DNA/adenovirus ASP2 vaccine to heart tissue after challenge.

CD8+ T lymphocytes play an important role in controlling infections by intracellular pathogens. Chemokines and their receptors are crucial for the migration of CD8+ T-lymphocytes, which are the main IFNγ producers and cytotoxic effectors cells. Although the participation of chemokine ligands and receptors has been largely explored in viral infection, much less is known in infection by Trypanosoma cruzi, the causative agent of Chagas disease. After T. cruzi infection, CXCR3 chemokine receptor is highly expressed on the surface of CD8+ T-lymphocytes. Here, we hypothesized that CXCR3 is a key molecule for migration of parasite-specific CD8+ T-cells towards infected tissues, where they may play their effector activities. Using a model of induction of resistance to highly susceptible A/Sn mice using an ASP2-carrying DNA/adenovirus prime-boost strategy, we showed that CXCR3 expression was upregulated on CD8+ T-cells, which selectively migrated towards its ligands CXCL9 and CXCL10. Anti-CXCR3 administration reversed the vaccine-induced resistance to T. cruzi infection in a way associated with hampered cytotoxic activity and increased proapoptotic markers on the H2KK-restricted TEWETGQI-specific CD8+ T-cells. Furthermore, CXCR3 receptor critically guided TEWETGQI-specific effector CD8+ T-cells to the infected heart tissue that express CXCL9 and CXCL10. Overall, our study pointed CXCR3 and its ligands as key molecules to drive T. cruzi-specific effector CD8+ T-cells into the infected heart tissue. The unveiling of the process driving cell migration and colonization of infected tissues by pathogen-specific effector T-cells is a crucial requirement to the development of vaccine strategies.

LFA-1 Mediates Cytotoxicity and Tissue Migration of Specific CD8+ T Cells after Heterologous Prime-Boost Vaccination against Trypanosoma cruzi Infection.

Integrins mediate the lymphocyte migration into an infected tissue, and these cells are essential for controlling the multiplication of many intracellular parasites such as Trypanosoma cruzi, the causative agent of Chagas disease. Here, we explore LFA-1 and VLA-4 roles in the migration of specific CD8+ T cells generated by heterologous prime-boost immunization during experimental infection with T. cruzi. To this end, vaccinated mice were treated with monoclonal anti-LFA-1 and/or anti-VLA-4 to block these molecules. After anti-LFA-1, but not anti-VLA-4 treatment, all vaccinated mice displayed increased blood and tissue parasitemia, and quickly succumbed to infection. In addition, there was an accumulation of specific CD8+ T cells in the spleen and lymph nodes and a decrease in the number of those cells, especially in the heart, suggesting that LFA-1 is important for the output of specific CD8+ T cells from secondary lymphoid organs into infected organs such as the heart. The treatment did not alter CD8+ T cell effector functions such as the production of pro-inflammatory cytokines and granzyme B, and maintained the proliferative capacity after treatment. However, the specific CD8+ T cell direct cytotoxicity was impaired after LFA-1 blockade. Also, these cells expressed higher levels of Fas/CD95 on the surface, suggesting that they are susceptible to programmed cell death by the extrinsic pathway. We conclude that LFA-1 plays an important role in the migration of specific CD8+ T cells and in the direct cytotoxicity of these cells.

Recombinant influenza A viruses harboring optimized dicistronic NA segment with an extended native 5' terminal sequence: induction of heterospecific B and T cell responses in mice.

We generated novel recombinant influenza A viruses (vNA38) harboring dicistronic NA segments with an extended native 5' terminal sequence of 70 nucleotides comprised of the last 42 nucleotides of the NA ORF and the 5' noncoding region (5' NCR). vNA38 viruses replicated stably and more efficiently than vNA35 viruses with a dicistronic NA segment comprised of the native 5' NCR only, that we described previously (Vieira Machado, A., Naffakh, N., van der Werf, S., Escriou, N., 2003. Expression of a foreign gene by stable recombinant influenza viruses harboring a dicistronic genomic segment with an internal promoter. Virology 313, 235-249). In addition, vNA38 viruses drove the expression of higher levels of encoded heterologous proteins than corresponding vNA35 viruses, both in cell culture and in the pulmonary tissue of infected mice. These data demonstrate that a sequence overlapping 5' coding and noncoding regions of the NA segment determines efficient replication and/or propagation of the vRNA. Intranasal immunization of mice with live vNA38 viruses induced B and T cell responses specific for the heterologous protein expressed, establishing the usefulness of such recombinant influenza viruses with a dicistronic segment for the development of live bivalent vaccines.