Amebic liver abscess (ALA), a parasitic disease due to infection with the protozoan Entamoeba histolytica, occurs age and gender dependent with strong preferences for adult males. Using a mouse model for ALA with a similar male bias for the disease, we have investigated the role of female and male sexual hormones and provide evidence for a strong contribution of testosterone. Removal of testosterone by orchiectomy significantly reduced sizes of abscesses in male mice, while substitution of testosterone increased development of ALA in female mice. Activation of natural killer T (NKT) cells, which are known to be important for the control of ALA, is influenced by testosterone. Specifically activated NKT cells isolated from female mice produce more IFNγ compared to NKT cells derived from male mice. This high level production of IFNγ in female derived NKT cells was inhibited by testosterone substitution, while the IFNγ production in male derived NKT cells was increased by orchiectomy. Gender dependent differences were not a result of differences in the total number of NKT cells, but a result of a higher activation potential for the CD4⁻ NKT cell subpopulation in female mice. Taken together, we conclude that the hormone status of the host, in particular the testosterone level, determines susceptibility to ALA at least in a mouse model of the disease.

The innate immune response is supposed to play an essential role in the control of amebic liver abscess (ALA), a severe form of invasive amoebiasis due to infection with the protozoan parasite Entamoeba histolytica. In a mouse model for the disease, we previously demonstrated that Jα18^-/- mice, lacking invariant natural killer T (iNKT) cells, suffer from more severe abscess development. Here we show that the specific activation of iNKT cells using α-galactosylceramide (α-GalCer) induces a significant reduction in the sizes of ALA lesions, whereas CD1d^−/− mice develop more severe abscesses. We identified a lipopeptidophosphoglycan from E. histolytica membranes (EhLPPG) as a possible natural NKT cell ligand and show that the purified phosphoinositol (PI) moiety of this molecule induces protective IFN-γ but not IL-4 production in NKT cells. The main component of EhLPPG responsible for NKT cell activation is a diacylated PI, (1-O-[(28∶0)-lyso-glycero-3-phosphatidyl-]2-O-(C16:0)-Ins). IFN-γ production by NKT cells requires the presence of CD1d and simultaneously TLR receptor signalling through MyD88 and secretion of IL-12. Similar to α-GalCer application, EhLPPG treatment significantly reduces the severity of ALA in ameba-infected mice. Our results suggest that EhLPPG is an amebic molecule that is important for the limitation of ALA development and may explain why the majority of E. histolytica-infected individuals do not develop amebic liver abscess. Amoebiasis is a widespread human parasitic disease caused by the intestinal protozoan Entamoeba histolytica. There are two major clinical manifestations of the disease, amebic colitis and amebic liver abscess. Interestingly, only a small proportion of E. histolytica-infected individuals develop invasive disease, whereas the majority harbors the parasite within the gut without clinical symptoms. So far, cells of the innate immune system have been described to constitute the main host defense mechanism for the control of amoebiasis, relying largely on the early production of interferon-γ (IFN-γ), a protein, which activates macrophages to kill microorganisms including parasites. However, the IFN-γ-producing cells as well as the amebic antigen involved in the activation have not been identified. Here we demonstrate that mice challenged with live E. histolytica, and which are deficient of a specific lymphocyte population known as natural killer T cells (NKT cells), have a reduced capacity to control ameba infection and develop much larger amebic liver abscesses compared to normal mice. In addition, we isolated a molecule from the surface membrane of E. histolytica, which constitutes a lipopeptidophosphoglycan, and which activates NKT cells for the production of protective IFN-γ. Thus, our study provides a mechanism for the innate control of ameba invasion that might explain why the majority of E. histolytica-infected individuals do not develop amebic disease.

Amebic liver abscess (ALA) is a focal destruction of liver tissue due to infection by the protozoan parasite Entamoeba histolytica (E. histolytica). Host tissue damage is attributed mainly to parasite pathogenicity factors, but massive early accumulation of mononuclear cells, including neutrophils, inflammatory monocytes and macrophages, at the site of infection raises the question of whether these cells also contribute to tissue damage. Using highly selective depletion strategies and cell-specific knockout mice, the relative contribution of innate immune cell populations to liver destruction during amebic infection was investigated. Neutrophils were not required for amebic infection nor did they appear to be substantially involved in tissue damage. In contrast, Kupffer cells and inflammatory monocytes contributed substantially to liver destruction during ALA, and tissue damage was mediated primarily by TNFα. These data indicate that besides direct antiparasitic drugs, modulating innate immune responses may potentially be beneficial in limiting ALA pathogenesis. Amebic liver abscess (ALA), an infectious disease caused by the protozoan parasite Entamoeba histolytica is characterized by severe focal liver damages. According to its name giving activity, destruction of liver tissue by E. histolytica has been attributed to parasite-specific effector molecules. However, abscess lesions contain considerable numbers of innate immune cells, such as neutrophils and macrophages, raising the question whether these host cells might contribute to the disease as well. We have investigated the role of the host immune response during ALA development using a mouse model for the disease. The results indicated that despite the presence of considerable numbers of neutrophils within the abscess lesions, these cells were dispensable for both the control of the disease and the tissue damage. On the other hand, two subsets of immune cells, the liver resident Kupffer cells and the inflammatory Ly6C-expressing monocytes were identified as the main effector cells responsible for liver tissue destruction. Furthermore, TNFα produced by the Ly6C-expressing monocytes, was found to be a cytokine that is critically involved in abscess development. Thus, our finding that host immune mechanisms are indeed responsible for liver tissue destruction during ALA development may change the view on the pathological mechanism of amebic disease.

Amebic liver abscess (ALA) is the most common extraintestinal manifestation of human infection by the enteric protozoan parasiteEntamoeba histolytica. In contrast to intestinal infection, ALA greatly predominates in males but is rare in females. Since humans are the only relevant host forE. histolytica, experimental studies concerning this sexual dimorphism have been hampered by the lack of a suitable animal model. By serial liver passage of culturedE. histolyticatrophozoites in gerbils and mice, we generated amebae which reproducibly induce ALA in C57BL/6 mice. Interestingly, all animals developed ALA, but the time courses of abscess formation differed significantly between the genders. Female mice were able to clear the infection within 3 days, whereas in male mice the parasite could be recovered for at least 14 days. Accordingly, male mice showed a prolonged time of recovery from ALA. Immunohistology of abscesses revealed that polymorphonuclear leukocytes and macrophages were the dominant infiltrates, but in addition, γ,δ-T cells, NK cells, and natural killer T (NKT) cells were also present at early times during abscess development, whereas conventional α,β-T cells appeared later, when female mice had already cleared the parasite. Interestingly, male and female mice differed in early cytokine production in response to ameba infection. Enzyme-linked immunospot assays performed with spleen cells of infected animals revealed significantly higher numbers of interleukin-4-producing cells in male mice but significantly higher numbers of gamma interferon (IFN-γ)-producing cells in female mice. Early IFN-γ production and the presence of functional NKT cells were found to be important for the control of hepatic amebiasis as application of an IFN-γ-neutralizing monoclonal antibody or the use of NKT knockout mice (Vα14iNKT, Jα 18^−/−) dramatically increased the size of ALA in female mice. In addition,E. histolyticatrophozoites could be reisolated from liver abscesses of Jα18^−/−mice on day 7 postinfection, when wild-type mice had already cleared the parasite. These data suggest that the sexual dimorphism in the control of ALA is due to gender-specific differences in early cytokine production mediated at least in part by NKT cells in response toE. histolyticainfection of the liver.

Hepatic amebiasis, predominantly occurring in men, is a focal destruction of the liver due to the invading protozoan Entamoeba histolytica. Classical monocytes as well as testosterone are identified to have important functions for the development of hepatic amebiasis in mice, but a link between testosterone and monocytes has not been identified. Here we show that testosterone treatment induces proinflammatory responses in human and mouse classical monocytes. When treated with 5α-dihydrotestosterone, a strong androgen receptor ligand, human classical monocytes increase CXCL1 production in the presence of Entamoeba histolytica antigens. Moreover, plasma testosterone levels of individuals undergoing transgender procedure correlate positively with the TNF and CXCL1 secretion from their cultured peripheral blood mononuclear cells following lipopolysaccharide stimulation. Finally, testosterone substitution of castrated male mice increases the frequency of TNF/CXCL1-producing classical monocytes during hepatic amebiasis, supporting the hypothesis that the effects of androgens may contribute to an increased risk of developing monocyte-mediated pathologies.