The Influence of Parasite-derived Chemokines in Leishmaniasis

Title of Document: THE INFLUENCE OF PARASITE-DERIVED CHEMOKINES IN LEISHMANIASIS Sean Martin Conrad, Doctor of Philosophy, 2006 Directed By: Professor David M. Mosser Department of Cell Biology and Molecular Genetics Transgenic chemokine-secreting parasites were generated and used to actively recruit immune cells into Leishmania lesions. It was hypothesized that the chemokine induced cell migration would influence the magnitude and character of the immune response and thereby effect the outcome of disease. Two different transgenic chemokinesecreting parasites were engineered. One transgenic parasite secretes murine MCP-1, a CC chemokine primarily responsible for macrophage recruitment. The other transgenic parasite secretes murine IP-10, a CXC chemokine known to attract activated T-cells. Both transgenic parasites transcribed murine chemokine mRNA, translated murine chemokine protein, and infected and replicated inside resting peritoneal macrophages similar to wild-type parasites. However, the two transgenic parasites caused diverse phenotypes in infected mice. The MCP-1 secreting parasites caused little or no detectable lesions, while the IP-10 secreting parasites caused lesions that were significantly larger than the wild-type infected mice. The healing phenotype caused by MCP-1 secreting parasites was further analyzed. Infection of BALB/c, C57BL/6, or MCP-1 knockout (KO) mice with MCP-1 secreting parasites resulted in minimal lesion development compared to mice infected with wild-type parasites. MCP-1 secreting parasites caused substantial lesions with relatively high numbers of parasites in CCR2 KO mice indicating that the parasites are viable and healthy, and that the lack of lesion development is CCR2dependent. The enumeration of transgenic MCP-1 parasites in lesions demonstrated a significant reduction in parasite numbers, which coincided with an increase in CCR2-positive macrophage migration on day 7. CCR2-positive macrophages isolated from ears of mice infected with transgenic MCP-1 parasites contained virtually no parasites, whereas infection with wild-type parasites resulted in heavily-infected macrophages in lesions. The lack of parasite survival in mice infected with MCP-1 secreting parasites suggests that parasite-derived MCP-1 is recruiting a population of CCR2-positive macrophages to the lesion that efficiently kill Leishmania parasites. In-vitro studies revealed that optimal parasite killing required the recruitment of CCR2-positive macrophages followed by stimulation with a combination of both MCP-1 and IFN-γ. This work suggests that the parasite-derived MCP-1 can recruit a restrictive population of CCR2-positive macrophages into lesions that can be optimally stimulated by MCP-1 and IFN-γ to efficiently kill Leishmania parasites. THE INFLUENCE OF PARASITE-DERIVED CHEMOKINES IN LEISHMANIASIS