ovalbumin and Chagas-Disease

This study describes the synthesis of glycopeptides NHAc[βGal]-(Thr)2 -[αGalNAc]-(Thr)2 -[αGlcNAc]-(Thr)2 Gly-OVA (1-OVA) and NHAc[βGal-αGalNAc]-(Thr)3 -[αLacNAc]-(Thr)3 -Gly-OVA (2-OVA) as mimetics of both T. cruzi and tumor mucin glycoproteins. These glycopeptides were obtained by solid-phase synthesis, which involved the prior preparation of the protected glycosyl amino acids αGlcNAc-ThrOH (3), αGalNAc-ThrOH (4), βGal-ThrOH (5), αLacNAc-ThrOH (6), and βGal-αGalNAc-ThrOH (7) through glycosylation reactions. Immunizations of mice with glycopeptides 1-OVA and 2-OVA induced high antibody titers (1:16 000), as verified by ELISA tests, whereas flow cytometry assays showed the capacity of the obtained anti-glycopeptides 1-OVA and 2-OVA antibodies to recognize both T. cruzi and MCF-7 tumor cells. In addition, antisera induced by glycopeptides 1-OVA and 2-OVA were also able to inhibit T. cruzi fibroblast cell invasion (70 %) and to induce antibody-mediated cellular cytotoxicity (ADCC) against MCF-7 cells, with 50 % reduction of cell viability.

Topics: Amino Acid Sequence; Animals; Antibodies; Cell Line, Tumor; Cell Survival; Chagas Disease; Glycopeptides; Humans; Immunization; Mice; Mucins; Neoplasms; Ovalbumin; Trypanosoma cruzi

Infection with Trypanosoma cruzi results in the development of both type 1 and type 2 patterns of cytokine responses during acute and chronic stages of infection. To investigate the role of Th1 and Th2 subsets of CD4(+) T cells in determining the outcome of T. cruzi infection in mice, we have developed T. cruzi clones that express OVA and have used OVA-specific TCR-transgenic T cells to generate OVA-specific Th1 and Th2 cells. BALB/c mice receiving 10(7) OVA-specific Th1 cells and then challenged with OVA-expressing T. cruzi G-OVA.GPI showed significantly lower parasitemia and increased survival in comparison to mice that received no cells. In contrast, recipients of OVA-specific Th2 cells developed higher parasitemias, exhibited higher tissue parasitism and inflammation, and had higher mortality than recipients of Th1 cells after infection with T. cruzi G-OVA.GPI. Mice receiving a mixture of both Th1 and Th2 OVA-specific cells also were not protected from lethal challenge. The protective effect of the OVA-specific Th1 cells was OVA dependent as shown by the fact that transfer of OVA-specific Th1 or Th2 cells failed to alter the course of infection or disease in mice challenged with wild-type T. cruzi. Immunohistochemical analysis of OVA-specific Th1 and Th2 cells at 4, 15, and 30 days postinfection revealed the persistence and expansion of these cells in mice challenged with T. cruzi G-OVA.GPI but not in mice infected with wild-type T. cruzi. We conclude that transfer of Ag-specific Th1 cells but not Th2 cells protect mice from a lethal infection with T. cruzi.

Topics: Adoptive Transfer; Amino Acid Sequence; Animals; Cell Division; Cells, Cultured; Chagas Disease; Epitopes, T-Lymphocyte; Glycosylphosphatidylinositols; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Transgenic; Molecular Sequence Data; Ovalbumin; Plasmids; Th1 Cells; Th2 Cells; Trypanosoma cruzi

We have previously shown that splenic gammadelta T cells from young but not aged BALB/c mice possess suppressor activity in vivo and in vitro during the acute phase of Trypanosoma cruzi infection. The present work was undertaken to investigate the suppressor activity of gammadelta T cells from T. cruzi-infected euthymic or athymic mice and the role of the thymus in modulating non-adherent spleen cell suppressor activity during the acute phase of infection. Splenic gammadelta T cells from aged or athymic BALB/c mice reconstituted with total spleen cells or non-reconstituted did not exhibit suppressor activity when added to full allogeneic, mixed lymphocyte cultures. In contrast, splenic gammadelta T cells from young euthymic BALB/c mice showed suppressor activity in vitro. Thymectomy reduced the splenic gammadelta T cell suppressor activity of young BALB/c mice in a time-dependent manner, following a T. cruzi challenge. The continuous provision of thymocytes to aged mice, young thymectomized mice or total spleen cell-reconstituted athymic mice could re-establish the gammadelta T cell suppressor activity. Of particular significance was the observation that the depletion of gammadelta T cells during the acute phase of T. cruzi infection restored the capacity of these mice to mount a humoral immune response to a non-related antigen such as ovalbumin. These results indicate that gammadelta T cells of extrathymic origin cannot mediate suppression and that the thymus has a role in the regulation of suppression during the acute phase of T. cruzi infection.

Topics: Acute Disease; Aging; Animals; Antibody Formation; Chagas Disease; Immunity, Cellular; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Nude; Ovalbumin; Spleen; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Thymectomy; Thymus Gland; Trypanosoma cruzi

Control of macrophage parasiticidal function by treatment with recombinant cytokines or their neutralizing antibodies modifies the severity of experimental Trypanosoma cruzi infections. However, so far, no direct in vivo evidence has demonstrated changes in disease outcome after altering the initial ratios of parasite-specific IFN-gamma and IL-10/IL-4-secretor cells in secondary lymphoid organs. To this end, a population of predominantly CD4+ parasite-Ag-reactive, IL-4- and IL-10-secreting T lymphocytes derived from T. cruzi-immunized mice was adoptively transferred to naive recipients. Compared with cell responses from normal mice, spleen cells of uninfected recipients proliferated significantly to T. cruzi Ag and produced much greater amounts of IL-4 and IL-10; lower IFN-gamma levels and increased IL-4/IL-10 levels were induced by Con A stimulation. Recipient mice challenged with T. cruzi presented overwhelming tissue and blood parasitemia and early death, contrasting with typically resistant controls. Uninfected recipients did not exhibit tissue damage following cell transfer. No disease exacerbation occurred in recipients of OVA-reactive CD4+, IL-4/IL-10-secreting T lymphocytes stimulated with OVA at the start of infection. On Day 6 postinfection, not only spleen cells but also LN cells from infected recipients showed decreased production of IFN-gamma and augmented secretion of IL-4/IL-10 compared to cells from untransferred infected mice. The results indicate that an imbalance of Th cell populations leading to the predominance of secreted IL-4 and IL-10 at the start of infection and the concomitant down-regulation of IFN-gamma secretion reversed the host's resistance to T. cruzi. Moreover, transfer of anti-T. cruzi Th2-type cells most likely favored the in vivo expansion of parasite-specific host cells toward a Th2 phenotype.

Topics: Animals; Antigens, Protozoan; CD4-Positive T-Lymphocytes; Cells, Cultured; Chagas Disease; Disease Susceptibility; Epitopes; Immunity, Cellular; Immunization, Passive; Immunotherapy, Adoptive; Interferon-gamma; Interleukin-10; Interleukin-2; Interleukin-4; Lymph Nodes; Lymphocyte Activation; Macrophages; Male; Mice; Mice, Inbred A; Mice, Inbred BALB C; Mice, Inbred C57BL; Muscle, Smooth; Organ Specificity; Ovalbumin; Saponins; Spleen

Some in vitro and in vivo biological activities of an octadecapeptide derived from an 85-kDa surface protein of Trypanosoma cruzi trypomastigote were studied. The peptide coupled to a carrier protein induced the proliferative response of lymph node cells from mice immunized with various antigens. Moreover, sera from mice immunized with the coupled peptide were found to contain antibodies against a number of self and nonself antigens: fibronectin, bovine serum albumin, myosin, tetanus toxoid, ovalbumin, keyhole limpet hemocyanin, and DNA. These results are discussed in the context of Chagas' disease immunopathology.

Topics: Amino Acid Sequence; Animals; Chagas Disease; Immunization; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Oligopeptides; Ovalbumin; Protozoan Proteins; Trypanosoma cruzi

A radioimmunoassay (RIA) originally designed to measure antibody responses to Trypanosoma cruzi in mice was adapted for use in the immunodiagnosis of Chagas' disease in humans. The assay utilizes biotinylated antibodies and 3H-avidin as the tracer molecules, and has proven to be both safe and sensitive. Results using the RIA and those from direct agglutination and indirect fluorescent antibody tests were comparable in most cases. Using the RIA, we were able to discriminate between mice infected with T. cruzi and Trypanosoma rangeli. Also, sera from Leishmania-infected individuals do not have detectable levels of antibodies capable of binding to T. cruzi. Intact, fixed epimastigotes of T. cruzi are used as the detecting antigen in the RIA and give results comparable to those obtained with intact trypomastigotes.

Topics: Adult; Aged; Animals; Antibodies; Avidin; Biotin; Chagas Disease; Female; Fluorescent Antibody Technique; Humans; Mice; Mice, Inbred C57BL; Middle Aged; Ovalbumin; Radioimmunoassay; Trypanosoma cruzi

A solid phase radioimmunoassay (RIA) to measure parasite-specific antibody responses has been developed. The assay utilizes 3H-labeled avidin as the indicator molecule and represents a marked improvement in detection of specific antibodies to Trypanosoma cruzi over previously used RIA. Avidin is easily labeled to high efficiency (3.6 X 10(5) cpm/micrograms) under very mild conditions using N-succinimidyl-[2,3-3H]propionate, and the high efficiency of avidin-biotin binding coupled with the increased safety and longer half-life of the labeled reagent give this method advantage over currently used 125I-labeled Ab RIAs.

Topics: Animals; Antibody Formation; Antibody Specificity; Avidin; Chagas Disease; Female; Immunoglobulin G; Immunoglobulin M; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Ovalbumin; Rabbits; Radioimmunoassay; Trypanosoma cruzi

Topics: Animals; Chagas Disease; Female; Immunization; Immunoglobulin E; Immunoglobulin G; Male; Mice; Mice, Inbred Strains; Ovalbumin; Rats; Time Factors