Intragastric immunization of rats with Entamoeba histolytica trophozoites induces cecal mucosal IgE, eosinophilic infiltration, and type I hypersensitivity.

We have investigated the role of IgE in the local immunity of intestinal amebiasis, a parasitic infection known to induce specific antibody-forming cells (AFC) and IgA antibodies in rodents and humans. We found that intragastric immunization of rats with glutaraldehyde-fixed Entamoeba histolytica trophozoites significantly increased antiameba AFC in the Peyer's patches and spleen and that the lamina propria of the cecum from immunized animals was infiltrated by eosinophils armed with IgE antibodies. Morphometric analysis showed that IgE-containing cells and eosinophils were nearly three times more abundant in the cecum of immunized rats. Antigenic challenge with amebal lysates provoked an increase in the short-circuit current and in the transepithelial potential difference in Ussing-chambered cecum preparations from immunized rats. Although eosinophilia and the increase of IgE are common consequences of infection by parasitic worms, our results indicate that local immunity in intestinal amebiasis also involves IgE deposition, eosinophil infiltration, and type I hypersensitivity, which may explain some symptoms of amebic dysentery such as colic, abdominal tension, tenesmus, and bloody stools.

Immunodominant Entamoeba histolytica antigens recognized by serum and intestinal antibodies after local or systemic immunization of mice with glutaraldehyde fixed trophozoites.

We performed an immunoblot analysis of the main E. histolytica proteins recognized by immune sera and intestinal fluids of Balb/c mice immunized with glutaraldehyde fixed trophozoites (GFT) by intragastric, rectal and intraperitoneal routes, to determine if there were differences in the amebic antigens immunodominantly recognized at mucosal and systemic levels. The antigen patterns recognized by mice immunized via intraperitoneal and rectal routes were complex and similar suggesting that the immunization route (systemic or local), does not influence the recognition pattern elicited at mucosal or systemic levels. However, the number of amebic bands recognized after intragastric immunization was very low. The molecular weights of the principal amebic proteins recognized by serum antibodies were 150-130, 116, 104, 84, 56, 42, 18, and 16 kDa. The intestinal fluids of mice immunized via intraperitoneal and rectal routes contained antibodies that recognized five bands of 220-200, 150-134, 93-84, 43-41, and 16-14 kDa. These results suggest that there are differences in the number of immunodominant amebic antigens recognized at mucosal and systemic levels. Moreover we found that the bands of 150, 39 and 19 kDa. were mainly recognized by IgG, whereas the bands of 116, 93, and 16 were mainly recognized by IgM, indicating differences between the antigens immunodominantly recognized by serum antibodies from different isotypes.

Entamoeba histolytica: induction and isotype analysis of antibody producing cell responses in Peyer's patches and spleen after local and systemic immunization in male and female mice.

The purpose of this work was to determine if anti-amebic antibody producing cell responses could be elicited in Peyer's patches and spleen in mice locally or systemically immunized with glutaraldehyde-fixed trophozoites of Entamoeba histolytica (GFT). The animals were inoculated with either a single or four doses of GFT via intragastric, rectal, and intraperitoneal routes. The anti-amebic antibody producing cell responses were analyzed by a spot forming cell assay (ELISPOT). The kinetics of antibody response revealed that a single dose of GFT by any route evoked anti-amebic responses in Peyer's patches and spleen. Furthermore, antibody producing cells of the three major isotypes were produced in both Peyer's patches and spleen of the mice receiving four doses of GFT, by either local or systemic routes. Our results indicate that immunization with GFT can induce a considerable number of specific antibody producing cells, which seem to remain in the Peyer's patches. After rectal and intraperitoneal immunization, females produced higher anti-amebic responses than males. Since either local or systemic immunization with GFT elicits both mucosal and systemic anti-amebic responses, this strategy should be considered as a promising tool for future elaboration of an anti-amebic vaccine.

The use of an ELISPOT assay to evaluate intestinal and systemic antibody responses to locally administered Entamoeba histolytica antigen in mice.

The local induction of humoral immune response to Entamoeba histolytica trophozoites in the gut has not been studied. This work reports some of our recent studies aimed to induce optimal immune responses against E. histolytica in mice and to describe a novel approach for monitoring mucosal immune responses induced in the gastrointestinal tract and expressed locally and systemically. We have compared the kinetics of both mucosal and systemic primary antibody responses to E. histolytica in the Peyer's patches (PP) and the spleen in Balb/c mice after a single dose of glutaraldehyde fixed amebas (GFA) by intragastric (IG), rectal (R), and intraperitoneal (IP) routes. The number of antibody-secreting cells directed to E. histolytica was assessed by the technique of ELISPOT on nitrocellulose filters. The antibody response to E. histolytica was detected in both PP and spleen with the three routes, indicating that either mucosal or systemic stimulation by GFA generates both types of response. We also determined the total antibody response in intestinal fluids and the antibody secretions from spleen and PP cells in vitro and found differences between male and female mice.

Interaction of cadmium with actin microfilaments in vitro.

Cadmium, an environmental and occupational health hazard, can substitute for calcium in the activation of calmodulin in several in vitro assays. It was shown that Cd(2+) can substitute for Ca(2+) in the induction of actin-based gelation in cytoplasmic extracts from rat liver; gelation induced by either Ca(2+) or Cd(2+) is inhibited by trifluoperazine, a well known calmodulin inhibitor; and in MDCK cells there is a Cd(2+)-induced redistribution of actin filaments with the loss of stress fibres and the appearance of actin bundles at the periphery of these cells.