1 Department of Gastroenterology, 2 Institute of Pathology, and 3 Department of Experimental Immunology, University Hospital of Basel, 4031 Basel, Switzerland, 4 Operative Unit of Pathology and Cytopathology, Regional Cancer Center, 85028 Rionero in Vulture, Italy
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Tumor necrosis
factor- (TNF-
) and interferon-
(IFN-
) are important for the
pathogenesis of Helicobacter pylori-associated gastritis and
peptic ulcer disease. Gastric biopsies from H. pylori-positive and -negative patients were used to examine the in
situ correlation of TNF-
and IFN-
with epithelial cell
apoptosis, bacterial load, and histological parameters of
gastritis. From the same patients, we isolated H. pylori-specific T cell lines and clones and examined their ex vivo
release of proinflammatory cytokines. We found a highly significant
correlation of TNF-
and IFN-
production with activity and grade
of gastritis (P < 0.01), H. pylori density (P = 0.01), epithelial cell apoptosis
(P < 0.001), and Fas/Fas-ligand expression
(P < 0.001). T cell lines and clones were all
TCR-
+ and showed T helper 1 functional phenotype.
With the use of serial histological sections, this study showed for the
first time the in situ correlation of TNF-
and IFN-
with
epithelial cell apoptosis, bacterial load, and histological
severity of disease and emphasizes the role of these cytokines in the
pathophysiology of H. pylori-associated disease.
tumor necrosis factor-; interferon-
; Fas, Fas-ligand; human
leukocyte antigen D-related
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
HELICOBACTER PYLORI INFECTION is the main cause of chronic gastritis and plays an important role in the pathogenesis of peptic ulcer disease, maltoma, and gastric cancer. The infection induces a chronic active inflammation with infiltrating neutrophils, T lymphocytes, B cells, and plasma cells. It has been postulated that differences of specific H. pylori strains as well as environmental and genetic factors of the host may determine the clinical outcome of H. pylori infection (5, 8, 22). Autoimmune-like reactions or the bystander effect associated with chronic active inflammation may explain the pathogenesis of peptic ulcer disease (12).
H. pylori infection generally persists for life with a widespread gastric inflammatory response, implying that the local mechanisms are ineffective in clearing the infection. Failure to resolve the infection probably results from induction of an inappropriate immune response and might be facilitated by an exaggerated activation of T cells.
In vitro studies suggest that in H. pylori infection, T
cells infiltrating the gastric mucosa mainly produce proinflammatory cytokines, such as tumor necrosis factor- (TNF-
) and
interferon-
(IFN-
). In previous studies, we examined the effect
of TNF-
and IFN-
on gastrin release (20) as well as
on migration and proliferation of isolated gastric cells
(18) and suggested that TNF-
could be one of the key
cytokines in H. pylori-associated gastritis and peptic ulcer
disease (20). Recently, several studies have been
published about the origin of TNF-
- and IFN-
-producing cells
(1, 21). Fas and Fas-ligand (Fas-L) expression have been
described in freshly isolated gastric epitelial cells, on mononuclear
cells, and in gastric T cell lines, as well as in gastric
adenocarcinoma (3). Furthermore, studies have shown the
association among H. pylori gastritis, T cell infiltration, and class II major histocompatibility complex expression in gastric epitelial cells (29, 33).
Cell proliferation and apoptosis are essential events for
normal cellular turnover of gastric tissue. In the healthy stomach, a
balance exists between proliferation of new epithelial cells and death
of senescent cells (15, 17). Induction of
apoptosis by H. pylori has been observed in patients
with gastritis and duodenal ulcers (15) and subsides after
successful H. pylori eradication (15, 17, 23,
25). Sustained stimulation of apoptosis could ultimately
result in excessive cell loss and ulcer development (in the acute
phase) or atrophy (in the chronic phase) and promote the subsequent
occurrence of gastric adenocarcinoma. The mechanism(s) by which
H. pylori induces apoptosis is unknown. TNF- and
IFN-
could be involved in the development of apoptosis as
well as signaling pathways including stimulation of specific cell death
receptors such as Fas/Fas-L. Fas expression has been described in
gastric carcinoma cells as well as in tissue samples of H. pylori-infected patients (26).
The aim of this study was to examine the in situ correlation of TNF-
and IFN-
with epithelial cell apoptosis, bacterial load, and
histological parameters of gastritis by the use of serial histological
sections from the same biopsies. From the same patients, we isolated
H. pylori-specific T cell lines and clones and investigated their ex vivo release of proinflammatory cytokines.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Patients and biopsies. Twenty H. pylori-positive patients and 12 H. pylori-negative controls were included in the study. H. pylori-positive patients had a median age of 57.5 (range 32-83) yr. Four patients had peptic ulcer disease, and 16 had dyspepsia. Patients treated with nonsteroidal anti-inflammatory drugs, antibiotics, proton pump inhibitors, H2 receptor antagonists, or bismuth at least 2 mo before study enrollment were excluded. H. pylori status was determined in all patients by urease test, histology, and culture. Patients were considered positive if at least two of three tests were positive. All patients with positive culture or immunohistochemistry were H. pylori positive. The H. pylori-negative control group consisted of 12 dyspeptic patients with a median age of 54 (range 26-82) yr.
Endoscopy. Upper gastrointestinal endoscopy was performed after overnight fast. Six antrum and three body biopsies were taken for histology, urease test, and bacterial cultures. Two antrum and one body specimens were placed in formalin and embedded in paraffin for histopathological examination. Two antrum and one body biopsies were snap frozen for immunohistochemical characterization of cytokine production and dUTP nick-end labeling (TUNEL) assay. One antrum biopsy was used for the urease test; one antrum and one body specimen were put in cell culture medium.
Histological and immunohistochemical assessment. Paraffin sections were stained with hematoxylin-eosin and modified Giemsa. Each biopsy specimen was assessed for the presence, type, and density of the inflammatory infiltrate and the presence of H. pylori. Gastritis was graded as absent, mild, moderate, and severe according to the updated Sydney system (10).
Characterization of inflammatory cells was performed by immunohistochemical staining with the ABC method according to the manufacturer's instructions (Vectastain Elite, ABC kit). The following antibodies were used for immunohistological studies: anti-CD20 (clone L26), rabbit anti-human CD3 (model A452), anti-CD68 (KP1) (all from DAKO, Carpintera, CA), anti-CD14 (Coulter), anti-human leukocyte antigen D-related (HLA-DR) (model L243; Dade-Behring, Düdingen, Switzerland), anti-TNF-Urease test. The urease test (Delta West, Perth, Australia) was performed according to the manufacturer's instructions. Results were considered positive if a definite red color developed within 24 h.
Bacteriologic assessment. H. pylori was cultured using two biopsies from the antral and body mucosa of each patient as previously described (11).
TUNEL assay. Apoptosis was examined by the terminal transferase-mediated biotinylated TUNEL method (TUNEL-kit FITC, Boeringher Mannheim, Mannheim, Germany). Frozen sections were thawed and treated according to the manufacturer's instructions. Slides were washed in PBS, then washed in H2O, mounted with Gelvatol-Gallat medium, and analyzed by fluorescence microscopy. At least 500 epithelial cells were counted in each section and the number of positive cells per 100 cells was expressed as the apoptotic index (in %).
T cell isolation and culture. T cells were cultured in RPMI 1640 medium supplemented with 2 mM Glutamax I, 100 µg/ml Kanamycin, 1 mM MEM nonessential amino acids, and 1 mM Na pyruvate (all from GIBCO-BRL, Life Technologies, Basel, Switzerland), 5% human serum (Blutspendezentrum, Basel, Switzerland), and 100 U of recombinant human interleukin-2 (IL-2). H. pylori-specific T cell lines were established from peripheral blood mononuclear cells (PBMC) by incubating 2 × 106 PBMC in IL-2-free RPMI medium and 100 µg/ml heat-inactivated H. pylori. After 5 days of culture, 10 U/ml IL-2 were added and the cultures were further expanded. After 2 wk, growing cells were restimulated using autologous-irradiated PBMC and 100 µg/ml heat-inactivated H. pylori. Specificity of T cells was assessed at days 28-32 of culture.
H. pylori-specific T cell lines were also raised from gastric biopsies as previously described (8). Briefly, biopsies were collected in complete RPMI medium added to gentamycin (50 µg/ml) (GIBCO-BRL), cyprofloxacyn (10 µg/ml) (Bayer, Zurich, Switzerland), and fungizone (2.5 µg/ml) (GIBCO-BRL), transferred in a petri dish and extensively washed to eliminate contaminating blood lymphocytes. Tissue was digested 5 h at 37°C with collagenase type IV (200 U/ml; Sigma, Buchs, Switzerland) and deoxyribonuclease type I (2 mg/ml; Sigma). Cells were washed and seeded at 1-5 × 105/ml together with the same amount of irradiated autologous PBMC as feeder cells and 100 µg/ml heat-inactivated H. pylori. T cell clones were established as previously described (8). Each clone was tested for reactivity against autologous Epstein-Barr virus (EBV)-transformed B cells in the presence and absence of heat-inactivated H. pylori using ELISA for detection of cytokines. Ag-specific clones were expanded, restimulated, and used in further experiments. From each donor, autologous EBV-transformed B cells were established as previously described (8).Flow cytometry of cultured T cells.
The following monoclonal antibodies (MAbs) were used: TR66 (anti-CD3),
B1 (anti-pan TCR-), anti-CD4, and anti-CD8 (Serotec, Oxford, UK) and FITC-labeled goat anti-mouse total Ig (Southern Biotechnology Associates, Birmingham, AL). Cells were stained as
reported (9). After washing, cells were immediately
analyzed using a FACScan flow cytometer (Becton Dickinson, San Jose,
CA). Dead cells were excluded by propidium iodide staining. Data
analysis was performed using the CELLQuest program (Becton Dickinson).
Cytokine release assays.
T cells (5 × 104/well) were stimulated with
heat-inactivated H. pylori (50 µg/ml) in triplicate
cultures in flat-bottom 96-well plates in the presence of
EBV-transformed autologous B cells as antigen-presenting cells (APCs;
5 × 104/well). After 18 h of incubation, 150 µl of supernatant were removed and used to test the content of
TNF-, IFN-
, and IL-4 by ELISA using commercial kits (R&D Systems,
Wiesbaden, Germany). Recombinant human cytokines were used as standard.
Statistical analysis.
A semiquantitative grading system (absent, minimal, moderate, and
severe) was adopted for immunohistochemical evaluation. Results were
further ranked into two grades: absent/minimal and moderate/severe. For
comparison of incidence among H. pylori-positive and
H. pylori-negative patients, 2-test or
Fisher's exact test were used, as appropriate. Furthermore, for
correlation of semiquantitative parameters a Spearman rank correlation
test was assessed as adequate. Calculations were performed running a
SAS software package on an Apple G3 computer.
P < 0.05 was considered significant.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Gastritis. In the H. pylori-positive group, four patients had mild, nine had moderate, and seven had severe chronic type B gastritis. In the H. pylori-negative control group, five had normal histology and seven had minimal lymphocytic infiltration of the tunica propria.
HLA-DR expression.
HLA-DR expression was observed in foveolar epithelium, superficial
stroma, and deep stroma. As suggested by Sarsfield et al. (28), HLA-DR expression was categorized as absent, scanty,
moderate, or abundant. Results were further ranked into two grades:
absent/scanty and moderate/abundant and a semiquantitative analysis was
performed. In all compartments, HLA-DR expression was significantly
more pronounced in H. pylori-infected samples
(P < 0.001; Table
1). In the foveolar epithelium,
HLA-DR expression was, in most cases, limited to the crypts and
virtually never occurred in the superficial epithelium. HLA-DR
expression was absent in intestinal metaplasia. In the stroma, the
majority of mononuclear cells expressed HLA-DR (Fig.
1, A and B). HLA-DR
expression was positively correlated with activity and grade of
gastritis as well as density of H. pylori (P < 0.001; Table 2).
|
|
|
In situ characterization of TNF-- and IFN-
-secreting cells.
Staining with anti-TNF-
MAb was exclusively detected in large
mononuclear cells, which were also stained with anti-CD68 and CD14 MAbs
(Fig. 1, C-E). In contrast, anti-IFN-
mAb
detected only T lymphocytes, identified by reactivity with anti-CD3
MAbs but not with anti-CD20 MAbs. The number of TNF-
- and
IFN-
-positive cells was significantly higher in H. pylori-positive biopsies (P < 0.01; Table 1).
TNF-
- (Fig. 1C) and IFN-
-positive cells showed a
diffuse distribution in the stroma of patients with moderate and severe
gastritis but were limited to the vicinity of gastric epithelium in
mild gastritis. In H. pylori-positive biopsies, the number
of TNF-
and IFN-
-expressing cells positively correlated with
activity and grade of gastritis (P < 0.05; Table 2)
and local density of H. pylori (P = 0.01;
Table 2). Cells stained with anti-IFN-
MAbs (i.e., recently
activated T cells) could exclusively be detected in H. pylori-positive patients.
Positive correlation of apoptosis with H. pylori gastritis. Apoptotic epithelial cells were observed in the superficial gastric mucosa of all patients. In H. pylori-positive samples, apoptotic cells were detected also in deep glands (Fig. 1H). The number of apoptotic epithelial cells was significantly higher in H. pylori-positive (P < 0.01; Table 1) (apoptotic index of 24.8, median 11.2, range 1-56%) than in H. pylori-negative biopsies (apoptotic index of 3.6, median 2.4, range 0-10%). The correlation of apoptosis and grade of gastritis was highly significant (P < 0.001; Table 2).
Fas/Fas-L expression. Fas/Fas-L expression was mainly detected in the epithelial cells of H. pylori-positive patients and only occasionally in stromal inflammatory cells (Fig. 1, F and G). Germinal centers in lymphoid follicles also showed dull Fas expression. The number of Fas-expressing epithelial cells was significantly higher in H. pylori-positive biopsies and in severe rather than mild gastritis (P < 0.05; Table 1). Fas-L expression was also more pronounced in H. pylori-positive samples (P < 0.001; Table 1). The correlation of Fas-L and activity/grade of gastritis was highly significant (P < 0.001; Table 2).
Correlation of cytokine production, apoptosis, and
Fas/Fas-L expression.
Apoptosis of stromal cells was significantly correlated with
the in situ detection of TNF- (P < 0.001) and
IFN-
(P < 0.05; Table 2). Apoptosis of
stromal cells was found in the vast majority of TNF-
-expressing
samples and only in one TNF-
-negative patient. Furthermore, TNF-
and IFN-
production was significantly correlated with Fas
(P < 0.05) and Fas-L expression (P < 0.001 for TNF-
; P < 0.05 for IFN-
).
Establishment of T cell lines and clones specific for H. pylori
from gastric biopsies.
To better characterize the nature of T lymphocytes infiltrating the
gastric mucosa of H. pylori-infected patients, T cell lines
were derived from antrum biopsies of 11 patients. Peripheral and
mucosal T cell lines showed the same reactivity. As control, 11 lines
were established from peripheral blood of the same donors. Ten lines
were derived from H. pylori-positive patients and one from a
negative donor. When the lines were tested in the presence of
EBV-transformed B cells as APCs and heat-inactivated H. pylori, 6 of 11 lines derived from gastric tissue showed in vitro
reactivity to H. pylori. The cell derived from the H. pylori-negative patient was nonreactive. Instead, 9 of 11 lines
from peripheral blood were H. pylori-reactive (Fig.
2). These T cells specifically released both TNF- and IFN-
on stimulation with H. pylori.
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
H. pylori infection is histologically characterized by
the presence of a large number of T cells in the lamina propria. A sizable fraction of these lymphocytes are specific for bacterial antigens and release proinflammatory lymphokines (32). Our
data confirm that in the gastric mucosa, the host response to H. pylori is mainly characterized by T lymphocytic infiltration and
local secretion of IFN- (21). In moderate and severe
gastritis, IFN-
-secreting T cells were diffuse in the stroma, likely
because they were locally stimulated. In these forms of gastritis, the
number of IFN-
-secreting T cells correlated with the density of
H. pylori. This finding suggests that in case of high
bacterial density, a large amount of antigen is available to
professional APCs that stimulate T cells in the stroma. In contrast, in
mild gastritis, IFN-
-secreting T cells were found in the proximity
of epithelial cells, indicating that in this form of gastritis, the
H. pylori stimulatory antigens remain mostly confined to
epithelial cells, where the bacteria are located. Perhaps, the same
epithelial cells present bacterial products to T cells, as shown in
vitro with other types of antigens (24). The finding that
a strong epithelial and stromal expression of HLA-DR molecules is
present during H. pylori infection supports this possibility.
HLA-DR expression, which we also found positively correlated with H. pylori density as well as with activity and grade of gastritis, is not homogeneous in the gastric mucosal epithelium. Isthmic areas were brightly stained, whereas the surface epithelium was dull as previously reported (26). This is a paradoxical finding, considering that epithelial cells are in intimate contact with mononuclear cells secreting proinflammatory cytokines and that these soluble factors induce the expression of high levels of HLA-DR on epithelial cells in vitro (30). It is tempting to speculate that mature epithelial cells migrating upward in the gastric mucosa become less sensitive to these cytokines. The low expression of human leucocyte antigen (HLA) class II molecules has no relevant consequences for antigen presentation, because T cells are mainly in contact with the epithelial cell lining at the bottom of crypts or in the isthmic regions, which instead are HLA class II bright. The absence of HLA-DR on superficial epithelial cells might instead be important to reduce bacterial adhesion, because H. pylori uses HLA class II molecules as additional cellular receptors (13).
Our data show that TNF- is released in the gastric mucosa during
H. pylori infection. With the use of immunohistochemistry, CD68-positive cells were stained with anti-TNF-
MAbs, whereas T
cells that appeared IFN-
positive were TNF-
-negative. This result
is different from what we observed in T cell clones. This difference
might be attributed to the lower sensitivity of immunohistochemistry versus ELISA for the detection of this cytokine, or to the presence of
high amounts of membrane-bound TNF-
on the surface of monocytes but
not of T cells in vivo. However, it is also possible that indeed
TNF-
is not produced by T cells when locally stimulated in the
gastric mucosa, and it is thus conceivable that clones do not exactly
reflect in vivo events. Further experiments are needed to determine
whether presentation of bacterial antigens by monocytes, dendritic
cells, and gastric epithelial cells stimulate secretion of different
lymphokines by T cells.
The finding that high numbers of locally activated T cells are present in patients with severe gastritis raises the question about the role of these cells in H. pylori infection. We agree with the current concept that H. pylori-specific T cells directly contribute to local inflammation.
H. pylori-specific T cell clones that we isolated from
gastric tissue showed a Th1-like functional phenotype. They released large amounts of TNF- and IFN-
in vitro, but no IL-4. T cell clones specific for H. pylori isolated from infected gastric
tissue have been previously studied, and they were predominantly Th1 as
well (6, 7). In addition, other investigators reported prevalence of Th1 phenotype in mitogen-activated polyclonal T cells
isolated from H. pylori-infected gastric mucosa
(2).
In our study, a second important observation was the detection of a large number of apoptotic cells in H. pylori-positive biopsies. Apoptotic epithelial cells were present both on the superficial mucosa and deep glands. In both locations, epithelial cells may have been killed by interaction with soluble cytotoxic products released by bacterial cells. This possibility is supported by the highly significant correlation that we found between apoptosis and number of H. pylori cells. Apoptosis was also significantly correlated with activity and grade of gastritis, which is at odds with other reports (17, 25). One possible explanation for this discrepancy is that previously published studies were performed on paraffin-embedded and not on frozen material, as in our study. We have already previously found that detection of apoptotic cells is much more sensitive in frozen samples than in paraffin-embedded material.
It is likely that multiple mechanisms cause apoptosis of
epithelial and stromal cells during H. pylori infection.
Local release of TNF- and IFN-
interaction between Fas and
Fas-L-expressing cells and direct killing by activated T cells or by
H. pylori cytotoxins may all contribute to the observed
apoptosis. TNF-
may induce cell apoptosis directly
(35) and can potentiate DNA-fragmentation induced by
H. pylori in vitro (34). It is remarkable that
among the parameters we studied, the presence of
TNF-
-secreting cells and apoptosis were those showing
the highest correlation (P < 0.001). Elevated and
persistent TNF-
secretion may contribute to the establishment of
tissue lesions reported in other inflammatory disease models
(19).
Like TNF-, IFN-
may also facilitate the H. pylori-induced apoptosis of epithelial cells. In the
literature, controversy exists whether local production of IFN-
is
increased in the gastric mucosa during H. pylori infection.
In vitro data suggest that IFN-
-producing T cells are present in
large numbers in infected mucosa (7). In our study,
IFN-
-secreting T lymphocytes were exclusively found in H. pylori-infected biopsies. Furthermore, a positive correlation was
detected between IFN-
expression and TUNEL-positive cells. IFN-
may modulate epithelial cell apoptosis by several mechanisms,
for example, by increasing Fas surface expression, by sensitizing
epithelial cells toward Fas-induced apoptosis, and by
upregulating TNF-
receptors, thus facilitating TNF-
-mediated
apoptosis (4, 16, 27).
Expression of Fas and Fas-L is a third mechanism that may directly contribute to apoptosis of epithelial cells. In contrast to another study (26), we found that epithelial cells and not stromal cells were Fas positive. Thus preferential expression of Fas by epithelial cells may explain why they were also the most frequent apoptotic cell type. We also found that epithelial cells express Fas-L.
Therefore, a cross talk among epithelial cells themselves might contribute to the development of the histological lesions. Similar findings have been reported in Hashimoto's thyroiditis, in which Fas/Fas-L interactions among thyrocytes lead to clinical hypothyroidism (14).
In conclusion, we found a positive correlation of TNF- and IFN-
with activity and grade of gastritis, H. pylori density, Fas/Fas-L expression, and apoptosis of epithelial cells. These findings support the important role of TNF-
and IFN-
in H. pylori-associated gastritis and peptic ulcer disease.
![]() |
ACKNOWLEDGEMENTS |
---|
This work was supported by Swiss National Fund Grants 3200-047662/1 and 31-045518.95 and by the Velux Foundation.
![]() |
FOOTNOTES |
---|
* F. S. Lehmann and L. Terracciano contributed equally to this work.
Address for reprint requests and other correspondence: F. S. Lehmann, Dept. of Gastroenterology, University Hospital of Basel, 4031 Basel, Switzerland.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
February 20, 2002;10.1152/ajpgi.00422.2001
Received 3 October 2001; accepted in final form 5 February 2002.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Ahlstedt, I,
Lindholm C,
Lonroth H,
Hamlet A,
Svennerholm AM,
and
Quiding-Jarbrink M.
Role of local cytokines in increased gastric expression of the secretory component in Helicobacter pylori infection.
Infect Immun
67:
4921-4925,
1999
2.
Bamford, KB,
Fan X,
Crowe SE,
Leary JF,
Gourley WK,
Luthra GK,
Brooks EG,
Graham DY,
Reyes VE,
and
Ernst PB.
Lymphocytes in the human gastric mucosa during Helicobacter pylori have a T helper cell 1 phenotype.
Gastroenterology
114:
482-492,
1998[ISI][Medline].
3.
Bennett, MW,
O'Connell J,
O'Sullivan GC,
Roche D,
Brady C,
Kelly J,
Collins JK,
and
Shanahan F.
Expression of Fas ligand by human gastric adenocarcinomas: a potential mechanism of immune escape in stomach cancer.
Gut
44:
156-162,
1999
4.
Bohm, W,
Thoma S,
Leithauser F,
Moller P,
Schirmbeck R,
and
Reimann J.
T cell-mediated, IFN-gamma-facilitated rejection of murine B16 melanomas.
J Immunol
161:
897-908,
1998
5.
Correa, P.
Helicobacter pylori and gastric carcinogenesis.
Am J Surg
1:
S37-S43,
1995.
6.
D'Elios, MM,
Manghetti M,
Almerigogna F,
Amedei A,
Costa F,
Burroni D,
Baldari CT,
Romagnani S,
Telford JL,
and
Del Prete G.
Different cytokine profile and antigen-specificity repertoire in Helicobacter pylori-specific T cell clones from the antrum of chronic gastritis patients with or without peptic ulcer.
Eur J Immunol
27:
1751-1755,
1997[ISI][Medline].
7.
D'Elios, MM,
Manghetti M,
De Carli M,
Costa F,
Baldari CT,
Burroni D,
Telford JL,
Romagnani S,
and
Del Prete G.
T helper 1 effector cells specific for Helicobacter pylori in the gastric antrum of patients with peptic ulcer disease.
J Immunol
158:
962-967,
1997[Abstract].
8.
De Libero, G.
Methods for the Generation of T Cell Clones and Epithelial Cell Lines from Excised Human Biopsies or Needle Aspirates. Oxford, UK: IRL, 1997.
9.
De Libero, G,
Rocci MP,
Casorati G,
Giachino C,
Oderda G,
Tavassoli K,
and
Migone N.
T cell receptor heterogeneity in gamma delta T cell clones from intestinal biopsies of patients with celiac disease.
Eur J Immunol
23:
499-504,
1993[ISI][Medline].
10.
Dixon, MF,
Genta RM,
Yardley JH,
and
Correa P.
Classification and grading of gastritis. The updated Sydney System International Workshop on the Histopathology of Gastritis, Houston 1994.
Am J Surg Pathol
20:
1161-1181,
1996[ISI][Medline].
11.
Engstrand, L,
Scheynius A,
Pahlson C,
Grimelius L,
Schwan A,
and
Gustavsson S.
Association of Campylobacter pylori with induced expression of class II transplantation antigens on gastric epithelial cells.
Infect Immun
57:
827-832,
1989[ISI][Medline];.
12.
Ernst, PB,
Jin Y,
Reyes VE,
and
Crowe SE.
The role of the local immune response in the pathogenesis of peptic ulcer formation.
Scand J Gastroenterol Suppl
205:
22-28,
1994[Medline].
13.
Fan, X,
Crowe SE,
Behar S,
Gunasena H,
Ye G,
Haeberle H,
Van Houten N,
Gourley WK,
Ernst PB,
and
Reyes VE.
The effect of class II major histocompatibility complex expression on adherence of Helicobacter pylori and induction of apoptosis in gastric epithelial cells: a mechanism for T helper cell type 1-mediated damage.
J Exp Med
187:
1659-1669,
1998
14.
Giordano, C,
Stassi G,
De Maria R,
Todaro M,
Richiusa P,
Papoff G,
Ruberti G,
Bagnasco M,
Testi R,
and
Galluzzo A.
Potential involvement of Fas and its ligand in the pathogenesis of Hashimoto's thyroiditis.
Science
275:
960-963,
1997
15.
Hahm, KB,
Lee KJ,
Choi SY,
Kim JH,
Cho SW,
Yim H,
Park SJ,
and
Chung MH.
Possibility of chemoprevention by the eradication of Helicobacter pylori: oxidative DNA damage and apoptosis in H. pylori infection.
Am J Gastroenterol
92:
1853-1857,
1997[ISI][Medline].
16.
Itoh, N,
Yonehara S,
Ishii A,
Yonehara M,
Mizushima S,
Sameshima M,
Hase A,
Seto Y,
and
Nagata S.
The polypeptide encoded by the cDNA for human cell surface antigen Fas can mediate apoptosis.
Cell
66:
233-243,
1991[ISI][Medline].
17.
Jones, NL,
Shannon PT,
Cutz E,
Yeger H,
and
Sherman PM.
Increase in proliferation and apoptosis of gastric epithelial cells early in the natural history of Helicobacter pylori infection.
Am J Pathol
151:
1695-1703,
1997[Abstract].
18.
Kato, K,
Chen MC,
Nguyen M,
Lehmann FS,
Podolsky DK,
and
Soll AH.
Effects of growth factors and trefoil peptides on migration and replication in primary oxyntic cultures.
Am J Physiol Gastrointest Liver Physiol
276:
G1105-G1116,
1999
19.
Keffer, J,
Probert L,
Cazlaris H,
Georgopoulos S,
Kaslaris E,
Kioussis D,
and
Kollias G.
Transgenic mice expressing human tumour necrosis factor: a predictive genetic model of arthritis.
EMBO J
10:
4025-4031,
1991[Abstract].
20.
Lehmann, FS,
Golodner EH,
Wang J,
Chen MC,
Avedian D,
Calam J,
Walsh JH,
Dubinett S,
and
Soll AH.
Mononuclear cells and cytokines stimulate gastrin release from canine antral cells in primary culture.
Am J Physiol Gastrointest Liver Physiol
270:
G783-G788,
1996
21.
Lindholm, C,
Quiding-Jarbrink M,
Lonroth H,
Hamlet A,
and
Svennerholm AM.
Local cytokine response in Helicobacter pylori-infected subjects.
Infect Immun
66:
5964-5971,
1998
22.
Malaty, HM,
Engstrand L,
Pedersen NL,
and
Graham DY.
Helicobacter pylori infection: genetic and environmental influences. A study of twins.
Ann Intern Med
120:
982-986,
1994
23.
Mannick, EE,
Bravo LE,
Zarama G,
Realpe JL,
Zhang XJ,
Ruiz B,
Fontham ET,
Mera R,
Miller MJ,
and
Correa P.
Inducible nitric oxide synthase, nitrotyrosine, and apoptosis in Helicobacter pylori gastritis: effect of antibiotics and antioxidants.
Cancer Res
56:
3238-3243,
1996[Abstract].
24.
Mayer, L,
and
Shlien R.
Evidence for function of Ia molecules on gut epithelial cells in man.
J Exp Med
166:
1471-1483,
1987[Abstract].
25.
Moss, SF,
Calam J,
Agarwal B,
Wang S,
and
Holt PR.
Induction of gastric epithelial apoptosis by Helicobacter pylori.
Gut
38:
498-501,
1996[Abstract].
26.
Rudi, J,
Kuck D,
Strand S,
von Herbay A,
Mariani SM,
Krammer PH,
Galle PR,
and
Stremmel W.
Involvement of the CD95 (APO-1/Fas) receptor and ligand system in Helicobacter pylori-induced gastric epithelial apoptosis.
J Clin Invest
102:
1506-1514,
1998
27.
Ruggiero, V,
Tavernier J,
Fiers W,
and
Baglioni C.
Induction of the synthesis of tumor necrosis factor receptors by interferon-.
J Immunol
136:
2445-2450,
1986
28.
Sarsfield, P,
Jones DB,
Wotherspoon AC,
Harvard T,
and
Wright DH.
A study of accessory cells in the acquired lymphoid tissue of Helicobacter gastritis.
J Pathol
180:
18-25,
1996[ISI][Medline].
29.
Scheynius, A,
and
Engstrand L.
Gastric epithelial cells in Helicobacter pylori-associated gastritis express HLA-DR but not ICAM-1.
Scand J Immunol
33:
237-241,
1991[ISI][Medline].
30.
Spencer, J,
Finn T,
and
Isaacson PG.
Expression of HLA-DR antigens on epithelium associated with lymphoid tissue in the human gastrointestinal tract.
Gut
27:
153-157,
1986[Abstract].
31.
Sternberger, LA,
Hardy PH, Jr,
Cuculis JJ,
and
Meyer HG.
The unlabeled antibody enzyme method of immunohistochemistry: preparation and properties of soluble antigen-antibody complex (horseradish peroxidase-antihorseradish peroxidase) and its use in identification of spirochetes.
J Histochem Cytochem
18:
315-333,
1970[ISI][Medline].
32.
Telford, JL,
Covacci A,
Rappuoli R,
and
Chiara P.
Immunobiology of Helicobacter pylori infection.
Curr Opin Immunol
9:
498-503,
1997[ISI][Medline].
33.
Valnes, K,
Huitfeldt HS,
and
Brandtzaeg P.
Relation between T cell number and epithelial HLA class II expression quantified by image analysis in normal and inflamed human gastric mucosa.
Gut
31:
647-652,
1990[Abstract].
34.
Wagner, S,
Beil W,
Westermann J,
Logan RP,
Bock CT,
Trautwein C,
Bleck JS,
and
Manns MP.
Regulation of gastric epithelial cell growth by Helicobacter pylori: offdence for a major role of apoptosis.
Gastroenterology
113:
1836-1847,
1997[ISI][Medline].
35.
Zheng, L,
Fisher G,
Miller RE,
Peschon J,
Lynch DH,
and
Lenardo MJ.
Induction of apoptosis in mature T cells by tumour necrosis factor.
Nature
377:
348-351,
1995[ISI][Medline].