7 research outputs found
Lactobacillus johnsonii La1 Attenuates Helicobacter pylori-Associated Gastritis and Reduces Levels of Proinflammatory Chemokines in C57BL/6 Mice
In clinical settings, Lactobacillus johnsonii La1 administration has been reported to have a favorable effect on Helicobacter pylori-associated gastritis, although the mechanism remains unclear. We administered, continuously through the water supply, live La1 to H. pylori-infected C57BL/6 mice and followed colonization, the development of H. pylori-associated gastritis in the lamina propria, and the levels of proinflammatory chemokines macrophage inflammatory protein 2 (MIP-2) and keratinocyte-derived cytokine (KC) in the serum and gastric tissue over a period of 3 months. We documented a significant attenuation in both lymphocytic (P = 0.038) and neutrophilic (P = 0.003) inflammatory infiltration in the lamina propria as well as in the circulating levels of anti-H. pylori immunoglobulin G antibodies (P = 0.003), although we did not observe a suppressive effect of La1 on H. pylori colonizing numbers. Other lactobacilli, such as L. amylovorus DCE 471 and L. acidophilus IBB 801, did not attenuate H. pylori-associated gastritis to the same extent. MIP-2 serum levels were distinctly reduced during the early stages of H. pylori infection in the La1-treated animals, as were gastric mucosal levels of MIP-2 and KC. Finally, we also observed a significant reduction (P = 0.046) in H. pylori-induced interleukin-8 secretion by human adenocarcinoma AGS cells in vitro in the presence of neutralized (pH 6.8) La1 spent culture supernatants, without concomitant loss of H. pylori viability. These observations suggest that during the early infection stages, administration of La1 can attenuate H. pylori-induced gastritis in vivo, possibly by reducing proinflammatory chemotactic signals responsible for the recruitment of lymphocytes and neutrophils in the lamina propria
Helicobacter pylori isolates from Greek children express type 2 and type 1 Lewis and alpha 1,6-glucan antigens in conjunction with a functional type IV secretion system
Helicobacter pylon infection is often acquired in childhood and can
persist for life. Previous studies in adult patients have shown that H.
pylori isolates from North American and European hosts express
predominantly type 2 Lewis x (Le(x)) and Le(y) epitopes, while Asian
strains have the capacity to express type 1 Le(a) and Le(b) structures.
In order to understand the influence of environmental and host factors
on the expression of Le antigens, we analysed 50 Greek H. pylori
isolates from symptomatic children. Both CagA-positive and -negative
strains were evaluated. The expression of Le antigens was determined by
whole-cell indirect ELISA (WCE), and LPS profiles were assessed by gel
electrophoresis and immunoblotting. Occurrence of Le(x) and/or Le(y)
antigens was confirmed in 35 of the isolates (70%) while 15 of the
isolates were non-typable. It was found that 11 of the paediatric
isolates had the propensity to express type 1 Le(b) blood-group antigen
(22%), a feature relatively uncommon in H. pylori isolates from adults.
One strain expressed both Le(b) and Le(a) antigens. The majority of the
isolates (49/50, 98%) expressed alpha 1,6-glucan, an antigenic non-Le
determinant present in the outer core region of H. pylon LPS. All Le(x)-
and Le(y)-expressing strains also carried a functional cag pathogenicity
island-encoding a type IV secretion system, capable of translocating
CagA protein, as well as the vacAs1 allele, suggesting that Le(x) and
Le(y) epitopes may aid the persistence of more aggressive strains. No
association between bacterial virulence characteristics and the
histopathological observations was evident
Strategy To Characterize the Number and Type of Repeating EPIYA Phosphorylation Motifs in the Carboxyl Terminus of CagA Protein in Helicobacter pylori Clinical Isolates
Cytotoxin-associated gene A (CagA) diversity with regard to EPIYA-A, -B, -C, or -D phosphorylation motifs may play an important role in Helicobacter pylori pathogenesis, and therefore determination of these motifs in H. pylori clinical isolates can become a useful prognostic tool. We propose a strategy for the accurate determination of CagA EPIYA motifs in clinical strains, based upon one-step PCR amplification using primers that flank the EPIYA coding region. We thus analyzed 135 H. pylori isolates derived from 75 adults and 60 children Greek patients. A total of 34 cases were found to be EPIYA PCR negative and were consequently verified as cagA negative by cagA-specific PCR, empty-site cagA PCR, and Western blotting. Sequencing of the remaining 101 PCR-positive amplicons confirmed that an accurate prediction of the number of EPIYA motifs on the basis of size distribution of the PCR products was feasible in all cases. Furthermore, our assay could identify closely related H. pylori subclones within the same patient, harboring different numbers of EPIYA repeats. The prevalence of CagA proteins with three EPIYA motifs (ABC) or four EPIYA motifs (ABCC) was the same within the adult and children groups. However, CagA species with more than four EPIYA motifs were observed exclusively within adults (8.6%), suggesting that CagA-positive strains may acquire additional EPIYA-C motifs throughout adulthood. Our strategy requires no initial cagA screening of the clinical isolates and can accurately predict the number of EPIYA repeats in single or multiple closely related subclones bearing different numbers of EPIYA motifs in their CagA, which may coexist within the same patient
Strategy to characterize the number and type of repeating EPIYA phosphorylation motifs in the carboxyl terminus of CagA protein in Helicobacter pylori clinical isolates
Cytotoxin-associated gene A (CagA) diversity with regard to EPIYA-A, -B,
-C, or -D phosphorylation motifs may play an important role in
Helicobacter pylori pathogenesis, and therefore determination of these
motifs in H. pylori clinical isolates can become a useful prognostic
tool. We propose a strategy for the accurate determination of CagA EPIYA
motifs in clinical strains, based upon one-step PCR amplification using
primers that Hank the EPIYA coding region. We thus analyzed 135 H.
pylori isolates derived from 75 adults and 60 children Greek patients. A
total of 34 cases were found to be EPIYA PCR negative and were
consequently verified as cagA negative by cagA-specific PCR, empty-site
cagA PCR, and Western blotting. Sequencing of the remaining 101
PCR-positive amplicons confirmed that an accurate prediction of the
number of EPIYA motifs on the basis of size distribution of the PCR
products was feasible in all cases. Furthermore, our assay could
identify closely related H. pylori subclones within the same patient,
harboring different numbers of EPIYA repeats. The prevalence of CagA
proteins with three EPIYA motifs (ABC) or four EPIYA motifs (ABCC) was
the same within the adult and children groups. However, CagA species
with more than four EPIYA motifs were observed exclusively within adults
(8.6%), suggesting that CagA-positive strains may acquire additional
EPIYA-C motifs throughout adulthood. Our strategy requires no initial
cagA screening of the clinical isolates and can accurately predict the
number of EPIYA repeats in single or multiple closely related subclones
bearing different numbers of EPIYA motifs in their CagA, which may
coexist within the same patient
CagA and VacA Polymorphisms Do Not Correlate with Severity of Histopathological Lesions in Helicobacter pylori-Infected Greek Children
The presence of various numbers of EPIYA tyrosine phosphorylation motifs
in the CagA protein of Helicobacter pylori has been suggested to
contribute to pathogenesis in adults. In this prospective study, we
characterized H. pylori isolates from symptomatic children, with
reference to the diversity of functional EPIYA motifs in the CagA
protein and vacA isotypes, and assessed the potential correlation with
the histopathological manifestations of the infection. We analyzed 105
H. pylori isolates from 98 children and determined the diversity of
EPIYA motifs in CagA by amplification and sequencing of the 3’ variable
region of the cagA gene as well as vacA isotypes for the signal, middle,
and intermediate regions. CagA phosphorylation and levels of secreted
IL-8 were determined following in vitro infection of AGS gastric
epithelial cells. Histopathological evaluation of H. pylori
colonization, activity, and severity of the associated gastritis was
performed according to the updated Sydney criteria. EPIYA A
(GLKN[ST]EPIYAKVNKKK), EPIYA B (Q[V/A]ASPEPIY[A/T]QV AKKVNAKI),
and EPIYA C (RS[V/A]SPEPIYATIDDLG) motifs were detected in the ABC
(46.6%) and ABCC (17.1%) combinations. No isolates harboring more than
two EPIYA C motifs in CagA were found. The presence of isogenic strains
with variable numbers of CagA EPIYA C motifs within the same patient was
detected in seven cases. Occurrence of increasing numbers of EPIYA C
motifs correlated strongly with presence of a high-vacuolation (s1 or
s2/i1/m1) phenotype and age. A weak positive correlation was observed
between vacuolating vacA genotypes and presence of nodular gastritis.
However, CagA- and VacA-dependent pathogenicities were not found to
contribute to severity of histopathology manifestations in H.
pylori-infected children