Role of tumor necrosis factor alpha in Helicobacter pylori gastritis in tumor necrosis factor receptor 1-deficient mice

Abstract

Increased gastric production of interleukin 8 and tumor necrosis factor alpha (TNF-alpha) has been implicated in the pathogenesis of Helicobacter pylori-associated gastroduodenal disease. In the present study we used a mouse model to demonstrate whether loss of the tumor necrosis factor receptor 1 (TNF-R1) function leads to differences in gastric inflammation or the systemic immune response in H. pylori infection. Six different clinical isolates of H. pylori (three cytotoxin-positive and three cytotoxin-negative strains) were adapted to C57BL/6 mice. TNF-R1-deficient (TNF-R1(-/-)) mice (n = 19) and isogenetic controls (n = 24) were infected and sacrificed after 4 weeks of infection. Inflammation of the stomach and the humoral immune response to H. pylori were evaluated by histological, immunohistochemical, and serological methods. There was no detectable difference in the grade or activity of gastritis in TNF-R1(-/-) mice when they were compared with wild-type mice, but the number of lymphoid aggregates was slightly reduced in the gastric mucosa of TNF-R1(-/-) mice. Interestingly, total immunoglobulin G (IgG), as well as IgG1, IgG2b, and IgG3, H. pylori-specific antibody titers were significantly higher in wild-type mice. As revealed by immunoblot analysis, the difference in reactivity against H. pylori antigens was not based on a failure to recognize single H. pylori antigens in TNF-R1(-/-) mice. We therefore suggest that TNF-R1-mediated TNF-alpha signals might support a systemic humoral immune response against H. pylori and that the gastric inflammatory response to H. pylori infection seems to be independent of TNF-R1-mediated signals.

FIG. 1.

Box plots with medians, minimum and maximum values, upper and lower quartiles, and extremes (asterisks) of IgG antibody titers (ELISA units) to H. pylori (Hp) in infected TNF-R1-deficient mice (TNF-R1^−/−, n = 19) and controls (TNF-R1^+/+, n = 24). H. pylori-specific IgG antibody titers were significantly higher in infected isogenetic control mice than in TNF-R1-deficient mice (P = 0.0049).

FIG. 2.

Box plots with medians, minimum and maximum values, upper and lower quartiles, outliers (○), and extremes (asterisks) of IgG subclass antibody responses (optical density [OD] in ELISA analysis) to H. pylori in infected TNF-R1-deficient mice (TNF-R1^−/−, n = 19) and controls (TNF-R1^+/+, n = 24). Significantly higher titers were found for IgG1 (open bars) (P = 0.0002), IgG2b (solid bars) (P < 0.0001), and IgG3 (bars with narrow cross-hatching) (P = 0.0008), but not for IgG2a (bars with broad cross-hatching) (P = 0.596), in infected isogenetic controls than in TNF-R1-deficient mice.

FIG. 3.

Positive reactions in immunoblots to individual antigens by infected TNF-R1-deficient mice (open bars) (n = 19) and isogenetic controls (solid bars) (n = 24). TNF-R1-deficient mice and wild-type mice did not differ significantly in reactivity against specific H. pylori antigens.

FIG. 4.

Mean values and standard deviations for number of cells per field of vision (magnification, ×400) for mice infected with cytotoxin-positive (solid bars) (n = 22) and cytotoxin-negative (shaded bars) (n = 21) H. pylori strains. Gastric mucosa in mice infected with cytotoxin-positive H. pylori strains showed significantly more polymorphonuclear leukocytes than gastric mucosa in mice infected with cytotoxin-negative H. pylori-strains (P < 0.0001). No differences was detected for any other cell type analyzed.

FIG. 5.

Infiltration of gastric mucosa with polymorphonuclear granulocytes (stained with MAb Gr1) showed no significant difference between TNF-R1-deficient mice (A and B) and wild-type mice (C and D) (magnification, ×200). Uninfected controls consisting of wild-type mice (C) and TNF-R1-deficient mice (D) showed significantly less (P < 0.0001) infiltration of the gastric mucosa with polymorphonuclear granulocytes.