Effects of oral vaccination and immunomodulation by cholera toxin on experimental Helicobacter pylori infection, reinfection, and gastritis

Abstract

Therapeutic vaccination is an attractive strategy to control infection and disease caused by Helicobacter pylori. In mice infected with H. pylori we have studied the protective effect of oral immunization with an H. pylori lysate preparation given together with the mucosal adjuvant cholera toxin (CT), both against the initial infection and against a later reinfection challenge. We have also examined the effects of treatment with the CT adjuvant alone on H. pylori infection and reinfection. Specific immunization with lysate was found to result in a sixfold reduction of the extent (bacterial load) of the primary infection and also to provide similar levels of protection against reinfection. However, these effects were associated with severe postimmunization gastritis. In contrast, oral treatment with CT alone at the time of initial infection, while unable to suppress the initial infection, gave rise to a 20-fold reduction in bacterial load upon reinfection without causing any associated gastric inflammation. Both the infected animals that were specifically immunized and those that were treated with CT only displayed increased in vitro proliferative responses of mononuclear cells to H. pylori antigens. Antibody levels in response to H. pylori were on the other hand only marginally increased after treatment with CT, whereas they were markedly elevated after immunization with lysate plus CT, with a rise in both (Th2-driven) immunoglobulin G1 (IgG1) and, especially, (Th1-driven) IgG2a antibodies. The results illustrate the complex balance between protection and harmful inflammation after postinfection vaccination against H. pylori as studied in a mouse model.

FIG. 1.

Protective effect by therapeutic immunization with an H. pylori lysate preparation. Groups of mice were infected with H. pylori SS1 and either given CT alone at the time and again 1 week after the infection (hatched bar) or immunized four times with lysate plus CT after the establishment of infection (open bar). Infected untreated mice served as controls (filled bar). Results show geometric mean numbers (plus standard errors of the means) of bacteria cultured from the mouse stomach 6 weeks after start of infection. Mice immunized with lysate plus CT were protected significantly from a primary infection (P < 0.01). Results represent the composite of two consecutive experiments with eight mice per group, the results of which were so similar and overlapping that data were pooled for the graphic representation. The Mann-Whitney test was used for statistical analysis of differences between groups. ∗∗, P < 0.01 compared to the control group.

FIG. 2.

Treatment with CT or immunization with lysate plus CT during a primary infection leads to protection against reinfection. Groups of mice were infected with H. pylori SS1 and treated with CT or immunized with lysate plus CT as indicated in the legend for Fig. 1. The primary infection was eradicated after 6 weeks with antibiotics, and the animals were reinfected with H. pylori SS1. Reinfection controls were naive mice infected with H. pylori SS1. Results represent the composite of two consecutive experiments with eight mice per group, the results of which were so similar and overlapping that the data were pooled for the graphic representation. Statistical evaluation of differences between treatment groups was done by the Mann-Whitney test. ∗∗, P < 0.01; ∗, P < 0.04; n.s, P > 0.05.

FIG. 3.

Histopathology of mice infected with H. pylori. (A) Oxyntic mucosa of an infected mouse (6 to 8 weeks old) displaying normal morphology. Chief cells and parietal cells in the oxyntic mucosa can be seen (arrows). (B) Mucosa of a mouse treated with CT and examined after reinfection challenge, showing a normal morphology comparable to that of infected mice except for mild focal inflammation. (C) Mucosa of an infected mouse vaccinated with lysate plus CT showing massive lymphocyte infiltration and destruction of chief cells and parietal cells, which are replaced by undifferentiated mucus-secreting cells. (D) Mice immunized with lysate plus CT after eradication and reinfection showing severe inflammation but also the reappearance of some parietal cells (arrow).