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Infect. Immun. doi:10.1128/IAI.01217-07
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

TLR-9 dependent macrophage activation by Entamoeba histolytica DNA

Faculty of Medicine, Department of Microbiology and Infectious Diseases, University of Calgary Health Sciences Centre, 3330 Hospital Dr. NW, Calgary, Alberta, T2N 4N1, Canada. Department of Medicine and Center for GI Biology and Disease, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599

^* To whom correspondence should be addressed. Email: kchadee{at}ucalgary.ca.

	Abstract

Activation of the innate immune system by bacterial DNA and other invertebrate DNA represents a pathogen recognition mechanism. In this study we have investigated macrophage responses to DNA from the intestinal protozoan parasite Entamoeba histolytica. E. histolytica genomic DNA (E.h DNA) was purified from log phase trophozoites and tested with mouse macrophage cell line, RAW 264.7. RAW cells treated with E.h DNA demonstrated an increase TNF- mRNA and protein production. TNF- production was blocked by pre-treatment with chloroquine or monensin. In fact, an NF-B luciferase reporter assay in HEK cells transfected with human TLR9 demonstrated that E.h DNA signaled through TLR9 similar to CpG-ODN. Immunofluorescence confirmed NF-B activation in RAW cells, as seen by nuclear translocation of the p65 sub-unit. Western blot analysis demonstrated MAPK activation by E.h DNA. E.h DNA effects were abolished in MYD88-/- mice derived macrophages. In the context of disease, immunization with E.h DNA protected gerbils from an E. histolytica challenge infection. Take together, these results demonstrate that E.h DNA is recognized by TLR9 to activate macrophages and may provide an innate defense mechanism, characterized by the induction of the inflammatory mediator TNF-.