Cytokine expression in pediatric Helicobacter pylori infection

Helicobacter pylori infection is one of the most common gastrointestinal infections worldwide and almost invariably causes chronic gastritis in the infected host. A predominant Th1 profile has been demonstrated in H. pylori-infected mucosa from adults, but no previous study has evaluated in situ cytokine expression in children. We therefore examined expression of proinflammatory, anti-inflammatory, and regulatory cytokines by immunohistochemistry in cryopreserved antral biopsy specimens from 10 H. pylori-infected and 10 uninfected children and correlated expression of cytokines with histology scores. Concomitant expression of interleukin-8 (IL-8), gamma interferon (IFN-gamma), IL-4, transforming growth factor beta, and tumor necrosis factor alpha was seen in 8/10 H. pylori-infected cases and in 5/10 noninfected cases; all H. pylori-infected subjects showed staining for at least two of the cytokines. The proportion of epithelial cytokine-specific staining did not differ significantly between the groups, either in surface or glandular epithelium. Furthermore, no significant differences were noticed between intraepithelial or lamina propria lymphocyte staining in the groups. There was, however, a tendency of higher numbers of IFN-gamma- and IL-8-positive cells in the H. pylori-infected group. IFN-gamma and IL-8 lamina propria lymphocyte expression correlated significantly with antrum chronic inflammation, but there was no correlation between histology scores and epithelial cytokine expression. When the same techniques were used, the cytokine response appeared to be smaller in H. pylori-infected children than in adults, and there was no clear Th1 dominance. These results therefore suggest a different mucosal immunopathology in children. It remains to be determined whether the gastric immune response is downregulated in children with H. pylori infection and whether this is relevant to the outcome of infection.     

Differential stimulation of interleukin-12 (IL-12) and IL-10 by live and killed Helicobacter pylori in vitro and association of IL-12 production with gamma interferon-producing T cells in the human gastric mucosa.

The objective of these experiments was to examine the ability of Helicobacter pylori to stimulate interleukin-10 (IL-10) or IL-12 and select for either Th1 or Th2 cells. Gastric biopsy specimens were collected from patients who were categorized with respect to the presence of H. pylori and gastric disease as well as their age, gender, medications, and other factors. As Th1 and Th2 cells are selected by IL-12 and IL-10, respectively, biopsy specimens were screened for mRNA and protein for these cytokines. Although mRNA for IL-12 and IL-10 was detected in biopsy specimens obtained from both infected and uninfected patients, IL-12 protein predominated. Levels of IL-10 and IL-12 in gastric tissue did not change in response to infection. Moreover, gamma interferon (IFN-gamma)-producing T cells were found in both the infected and the uninfected gastric mucosa. Stimulation of peripheral blood leukocytes from either infected or uninfected donors with various concentrations of live or killed H. pylori induced immunoreactive IL-12 and IL-10. After stimulation with live H. pylori, IL-12 levels increased more than 30-fold, whereas IL-10 levels increased only 2- to 5-fold, compared to cells stimulated with medium alone. Interestingly, killed H. pylori induced significantly more IL-10 (P < 0.05) than live H. pylori, while recombinant urease only induced IL-10. These results demonstrate that live H. pylori selectively stimulates the induction of IL-12 and Th1 cells that produce IFN-gamma, whereas preparations used in oral vaccines induce more IL-10 and may favor Th2 cell responses.     

Helicobacter pylori-infected macrophages induce Th17 cell differentiation

Th17 cells represent a novel subset of CD4(+) T cells, which is associated with chronic inflammation. The present study evaluated Th17 cell responses to Helicobacter pylori infection in mouse model and CD4(+) T cell differentiation in response to H. pylori-infected macrophages. Th17 cells were observed in the H. pylori-infected gastric tissue. Co-culture of CD4(+) T cells with H. pylori-infected macrophages elevated IL-17 and IFN-γ secretion, up-regulated retinoid-related orphan receptor gamma t (RORγt) and T box expressed in T cells (T-bet) expression and increased the numbers of Th17 and Th1 cells. The expression of CD40, CD80, and CD86 and the secretion of IL-6, TGF-β1, IL-23, and CCL20 were significantly increased in H. pylori-stimulated macrophages. NF-κB pathway participated in the production of IL-6, IL-23, and CCL20 from macrophages in response to H. pylori, and inhibition of NF-κB pathway of macrophages resulted in less Th17 cell differentiation. Taken together, these results suggest that H. pylori induces Th17 cell differentiation via infected macrophages.     

STAT1 is essential for antimicrobial effector function but dispensable for gamma interferon production during Toxoplasma gondii infection

The opportunistic protozoan Toxoplasma gondii is a prototypic Th1-inducing pathogen inducing strong gamma interferon (IFN-gamma) cytokine responses that are required to survive infection. Intracellular signaling intermediate STAT1 mediates many effects of IFN-gamma and is implicated in activation of T-bet, a master regulator of Th1 differentiation. Here, we show that T. gondii-infected STAT1-null mice fail to upregulate the IFN-gamma-dependent effector molecules inducible nitric oxide synthase (iNOS), IGTP, and LRG-47, which are required for mice to survive infection. Both T-bet and interleukin-12 receptor beta2 (IL-12Rbeta2) failed to undergo normal upregulation in response to T. gondii. Development of IFN-gamma-producing CD4(+) and CD8(+) T lymphocytes was severely curtailed in the absence of STAT1, but a substantial level of STAT1-independent non-T-cell-derived IFN-gamma was induced. Absence of STAT1 also resulted in increased IL-4, Arg1, Ym1, and Fizz1, markers of Th2 differentiation and alternative macrophage activation. Together, the results show that T. gondii induces STAT1-dependent T-lymphocyte and STAT1-independent non-T-cell IFN-gamma production, but that effector functions of this type 1 cytokine cannot operate in the absence of STAT1, resulting in extreme susceptibility to acute infection.     

CCL28 is increased in human Helicobacter pylori-induced gastritis and mediates recruitment of gastric immunoglobulin A-secreting cells

Human Helicobacter pylori infection gives rise to an active chronic gastritis and is a major risk factor for the development of duodenal ulcer disease and gastric adenocarcinoma. The infection is accompanied by a large accumulation of immunoglobulin A (IgA)-secreting cells in the gastric mucosa, and following mucosal immunization only H. pylori-infected volunteers mounted a B-cell response in the gastric mucosa. To identify the signals for recruitment of gastric IgA-secreting cells, we investigated the gastric production of CCL28 (mucosa-associated epithelial chemokine) and CCL25 (thymus-expressed chemokine) in H. pylori-infected and uninfected individuals and the potential of gastric B-cell populations to migrate toward these chemokines. Gastric tissue from H. pylori-infected individuals contained significantly more CCL28 protein and mRNA than that from uninfected individuals, while CCL25 levels remained unchanged. Chemokine-induced migration of gastric lamina propria lymphocytes isolated from patients undergoing gastric resection was then assessed using the Transwell system. IgA-secreting cells and IgA(+) memory B cells from H. pylori-infected tissues migrated toward CCL28 but not CCL25, while the corresponding cells from uninfected patients did not. Furthermore, IgG-secreting cells from H. pylori-infected patients did not migrate to CCL28 but instead to CXCL12 (SDF-1alpha). However, chemokine receptor expression did not correlate to the migratory pattern of the different B-cell populations. These studies are the first to show increased CCL28 production during gastrointestinal infection in humans and provide an explanation for the large influx of IgA-secreting cells to the gastric mucosa in H. pylori-infected individuals.     

Instruction of naive CD4+ T-cell fate to T-bet expression and T helper 1 development: roles of T-cell receptor-mediated signals

Using T-cell receptor (TCR) transgenic mice, we demonstrate that TCR stimulation of naive CD4(+) T cells induces transient T-bet expression, interleukin (IL)-12 receptor beta2 up-regulation, and GATA-3 down-regulation, which leads to T helper (Th)1 differentiation even when the cells are stimulated with peptide-loaded I-A(b)-transfected Chinese hamster ovary cells in the absence of interferon-gamma (IFN-gamma) and IL-12. Sustained IFN-gamma and IL-12 stimulation augments naive T-cell differentiation into Th1 cells. Intriguingly, a significant Th1 response is observed even when T-bet(-/-) naive CD4(+) T cells are stimulated through TCR in the absence of IFN-gamma or IL-12. Stimulation of naive CD4(+) T cells in the absence of IFN-gamma or IL-12 with altered peptide ligand, whose avidity to the TCR is lower than that of original peptide, fails to up-regulate transient T-bet expression, sustains GATA-3 expression, and induces differentiation into Th2 cells. These results support the notion that direct interaction between TCR and peptide-loaded antigen-presenting cells, even in the absence of T-bet expression and costimulatory signals, primarily determine the fate of naive CD4(+) T cells to Th1 cells.