Magdalena Radwanska

Alternative Macrophage Activation Is Essential for Survival during Schistosomiasis and Downmodulates T Helper 1 Responses and Immunopathology

Macrophage/neutrophil-specific IL-4 receptor alpha-deficient mice (LysM(Cre)IL-4Ralpha(-/flox)) were generated to understand the role of IL-4/IL-13 responsive myeloid cells during Type 2 immune responses. LysM(Cre)IL-4Ralpha(-/flox) mice developed protective immunity against Nippostrongylus brasiliensis accompanied by T(H)2 development and goblet cell hyperplasia. In contrast, LysM(Cre)IL-4Ralpha(-/flox) mice were extremely susceptible to Schistosoma mansoni infection with 100% mortality during acute infection. Mortality was not dependent on neutrophils and occurred in the presence of T(H)2/Type 2 responses, granuloma formation, and egg-induced fibrosis. Death was associated with increased T(H)1 cytokines, hepatic and intestinal histopathology, increased NOS-2 activity, impaired egg expulsion, and sepsis. IL-10 was not able to compensate for the absence of IL-4/IL-13-activated alternative macrophages. Together, this shows that alternative macrophages are essential during schistosomiasis for protection against organ injury through downregulation of egg-induced inflammation.

Interferon-γ and Nitric Oxide in Combination with Antibodies Are Key Protective Host Immune Factors during Trypanosoma congolense Tc13 Infections

The control of chronic Trypanosoma congolense trypanosomiasis was analyzed using several gene-deficient mouse strains. First, interferon (IFN)-gamma receptor (IFN-gamma-R)-deficient mice were used to show that IFN- gamma -mediated immune activation is crucial for parasitemia control. Second, infections in major histocompatibility complex (MHC) class II-deficient mice indicate that this molecule is needed for initiation of IFN- gamma and subsequent tumor necrosis factor (TNF) production. Downstream of IFN-gamma-R signaling, inducible NO synthase (iNOS)-dependent trypanosome killing occurs, as is shown by the hypersusceptible phenotype of iNOS-deficient mice. Besides proinflammatory responses, B cells and, more specifically, immunoglobulin (Ig) G antibodies are crucial for parasite killing. Hence, parasitemia control is abolished in B cell-deficient mice, whereas IgM-deficient mice control the infection as efficiently as do wild-type mice. In addition, splenectomized mice that have a normal IgM response but an impaired IgG2a/3 response fail to control T. congolense infection. Collectively, these results suggest that host protective immunity against T. congolense is critically dependent on the combined action of the proinflammatory mediators/effectors IFN- gamma , TNF, and NO and antiparasite IgGs.

P75 Tumor Necrosis Factor–Receptor Shedding Occurs as a Protective Host Response during African Trypanosomiasis

In experimental murine trypanosomiasis, resistance is often scored as the capacity to control peak parasitemia levels, which results in prolonged survival. Infection-induced pathology has not systematically been used as a resistance criterion. Because this parameter could be the most relevant for comparative analysis of natural and experimental infections, as well as for understanding of pathology-associated immune alterations, we analyzed Trypanosoma brucei infections in 4 different established conventional mouse models, as well as in tumor necrosis factor (TNF)-deficient and TNF-receptor-deficient mice. Results indicate the following: (1) there is no correlation between peak parasitemia control or survival and the induction of infection-associated anemia, loss of body weight, liver pathology, reduced locomotor activity, and general morbidity; (2) serum levels of TNF, interferon- gamma, and interleukin-10, which are known to affect survival, do not correlate with induction of pathology; and (3) infection-induced occurrence of lipopolysaccharide hypersensitivity does not correlate with survival. However, one parameter that was found to correlate with the inhibition of trypanosomiasis-associated pathology in all models was the shedding of soluble p75 TNF-receptor during peak parasitemia stages. These results are important for future cytokine and trypanosomiasis pathology studies, because the interplay between TNF and the soluble receptors it sheds has not been considered in either human clinical sleeping sickness studies or in veterinary trypanosomiasis research.

Trypanosoma brucei infection elicits nitric oxide-dependent and nitric oxide-independent suppressive mechanisms.

During murine Trypanosoma brucei infection, macrophages contribute significantly to the inhibition of T cell responses. Although nitric oxide (NO) was shown to play a central role in macrophage‐mediated splenic suppression, macrophage‐mediated lymph node suppression occurred in an interferon‐γ (IFN‐γ)‐dependent manner. In this study, using NO inhibitor NG‐monomethyl‐L‐arginine and anti‐IFN‐γ antibodies, the relative contribution of NO and IFN‐γ to the active inhibition of ex vivo concanavalin A‐induced T cell proliferation taking place in the spleen and the lymph nodes of T. brucei‐infected mice was investigated. NO contributes to the suppressive activity of spleen and lymph node cells only during early‐stage infection. The existence of NO‐independent suppressive pathway was further evidenced in IFN‐γ‐/‐‐infected mice. Spleen cells from such animals do not produce NO but exert significant suppressive activity during the whole course of infection. In contrast in the lymph nodes, no suppressive activity is recorded at any moment of infection. Moreover, addition of exogenous IFN‐γ to cultures containing lymph node cells from IFN‐γ‐/‐‐infected mice does not impair proliferation despite NO production in such cultures. Thus during late‐stage infection, an IFN‐γ‐independent suppressive mechanism is elicited in the spleen, whereas in the lymph nodes, IFN‐γ is required yet not sufficient to inhibit T cell proliferation. J. Leukoc. Biol. 63: 429–439; 1998.

Tumor necrosis factor (TNF) receptor-1 (TNFp55) signal transduction and macrophage-derived soluble TNF are crucial for nitric oxide-mediated Trypanosoma congolense parasite killing.

Control of Trypanosoma congolense infections requires an early cell-mediated immune response. To unravel the role of tumor necrosis factor (TNF) in this process, 6 different T. congolense strains were used in 6 different gene-deficient mouse models that included TNF(-/-), TNF receptor-1 (TNFp55)(-/-), and TNF receptor-2 (TNFp75)(-/-) mice, 2 cell type-specific TNF(-/-) mice, as well as TNF-knock-in mice that expressed only membrane-bound TNF. Our results indicate that soluble TNF produced by macrophages/neutrophils and TNFp55 signaling are essential and sufficient to control parasitemia. The downstream mechanism in the control of T. congolense infection depends on inducible nitric oxide synthase activation in the liver. Such a role for nitric oxide is corroborated ex vivo, because the inhibitor N(G)-monomethyl-l-arginine blocks the trypanolytic activity of the adherent liver cell population, whereas exogenous interferon- gamma that stimulates nitric oxide production enhances parasite killing. In conclusion, the control of T. congolense infection depends on macrophage/neutrophil-derived soluble TNF and intact TNFp55 signaling, which induces trypanolytic nitric oxide.

Antibodies raised against the flagellar pocket fraction of Trypanosoma brucei preferentially recognize HSP60 in cDNA expression library.

A purified flagellar pocket fraction of the Trypanosoma brucei AnTat 1.1E clone was used for the generation of polyclonal antiserum in rats. Anti‐flagellar pocket antibodies present in this serum recognized several proteins distinct from the major variant surface glycoprotein (VSG). In Balb/c mice, flagellar pocket immunization resulted in partial resistance towards the challenge with a low dose of parasites. This was accompanied by the induction of specific IgG2a antibodies. In an attempt to discover protective parasite antigens, antiflagellar pocket serum was used for the screening of a T. brucei bloodstream form cDNA library constructed in the λgt11 bacteriophage expression system. Through antibody panning and VSG elimination, 15 specific cDNA inserts were selected. Most intriguing was the observation that in addition to two clones encoding the invariant surface glycoprotein 75 (ISG75), 10 out of 15 independently selected cDNA inserts encoded the trypanosome heat shock protein 60 (tHSP60).

A transcript encoding a proteasome beta-subunit and a zinc finger protein in Trypanosoma brucei brucei

During the screening of a Trypanosoma brucei brucei (T. b. brucei) cDNA library constructed from bloodstream form mRNA, we identified a 2.3kb cDNA encoding a proteasome beta subunit (ORF1) and a putative zinc finger protein (ORF2). Northern blot analysis indicated the presence of a digenic transcript as well as the two individual messengers in both procyclic and bloodstream forms of the parasite. Southern blot analysis showed the relevant locus to be unique. ORF1 encoded a 22.7kDa protein sharing over 50% identity with the eukaryotic PRCE (aka beta5) proteasome beta subunit. This protein contained a beta amino acid signature and residues involved in the catalytic activity. Further phylogenetic analysis indicated that this subunit as well as those from other kinetoplastids could be confidentially assigned to extant eukaryotic subfamilies such as beta1, beta2, and beta5. ORF2 encoded a 14.6kDa putative zinc finger protein containing five repeats of a CCHC motif commonly present in retroviral nucleocapsid proteins as well as proteins involved in vertebrate embryogenesis.

Trypanosomes and trypanosomiasis

Trypanosomes and trypanosomiasis / , Trypanosomes and trypanosomiasis / , کتابخانه دیجیتال جندی شاپور اهواز

Adaptive Immunity and Trypanosomiasis-Driven B-Cell Destruction

African Trypanosomiasis is an excellent model system to study immune escape by invading extracellular pathogens. Being under continuous attack by the host humoral response, trypanosomes developed a system of antigenic variation of their surface coat in order to evade antibody-mediated immune destruction. In-depth studies on the mechanisms of antigenic variation have resulted in the understanding of both structural and genetic aspects of the surface coat organization of trypanosomes, and the variant-specific glycoproteins (VSG) itself, i.e., the protein that provides the interface between the parasite and the host immune system (see Chaps. 1 and 3). To date, the current hypothesis of VSG-mediated antibody escape implies that during infection the host is capable of mounting an ever changing antibody repertoire, which allows to target in a specific manner each new trypanosome wave. In this chapter, a number of recent and new insights will be discussed that highlight the complexity of this system. Indeed, experimental data obtained in rodent T. brucei models suggest that anti-VSG responses are very short lived, and no effective memory is mounted during infection against the successive waves of occurring VATs. In addition, active destruction of the host B-cell compartment occurs during infection, affecting both trypanosome specific and non-specific B-cell memory. These finding will be discussed in the context of the long string of vaccine failure results that have hampered the initiation of an effective vaccine program for trypanosomiasis. Finally, this chapter will also provide new insights into antibody engineering that allow interfering with trypanosome biology in ways that are not part of the natural evolutionary pressure. Hence, possible new tools can be developed that can help in a sustainable long-term battle against both human and animal trypanosomiasis.