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SECRETED ENTAMOEBA HISTOLYTICA PRODUCTS FACILITATE PARASITE COLONIZATION IN THE COLON THROUGH INDUCTION OF IMMUNE TOLERANCE
JM Rybicka, K Chadee
University of Calgary, Calgary, Alberta
Amebiasis is estimated to currently affect approximately 10% of the world’s population; however in over 99% of cases the course of disease is completely asymptomatic and only a minority of patients develop amoebic colitis and/or liver abscess. The mechanisms that allow the parasite to be a successful colonizer in the absence of an inflammatory or immune response are not known. We hypothesized that products secreted in the colonic lumen by E. histolytica trophozoites may cause tolerance induction in antigen presenting cells, through upregulation and/or activation of inhibitory ILT2 receptor, which upon phosphorylation can block CD64 signaling and cell activation through recruitment of SHP-1, thus leading to immune suppression and facilitatating parasitic colonization. To test this hypothesis, murine macrophages or dendritic cells (DC) were cultured in the presence of splenic T lymphocytes and were exposed to 2 µg – 100 µg/mL of secreted (SP) and soluble (SAP) intrinsic amoebic proteins or LPS (2 µg/mL) as a control. As evidenced by FACS, SAP stimulation of macrophages resulted in a 71% decrease in the number of ILT2+CD3+CD4+ T cells as compared to unstimulated controls, while 10 µg/mL of SP, but not 10 µg/mL of SAP, caused an 8-fold increase in the population of ILT2 positive CD4+ T cells. Conversly, ILT2 expression in macrophages was not significantly affected by treatment with either SP or SAP; however, low concentrations of SP and SAP increased phosphorylation of the ILT2 receptor. Co-culturing of DC with T cells in the presence of 10 ug/mL of SP or SAP resulted in 3-fold increase in the number of ILT2 positive CD4+ T cells. Conversly, SP had no significant effect on ILT2 expression on DCs, while LPS cause a 23% decrease in ILT2 positive DCs and SAP a 33% increase. Furthermore, SP treatment increased in IL-10 mRNA production in both macrophages and DCs and promoted T cell apoptosis in macrophage presence. These findings suggest that E. histolytica secreted products can induce immune tolerance via upregulation of IL-10 production as well as activation of the inhibitory ILT2 receptor in macrophages and DCs leading to induction of apoptosis in T cells. Thus, E. histolytica causes deletion of the non-tolerogenic T cells and immunosupression, which allows the parasite to survive in the colon and avoid detection by scavenging immune cells.