[Show abstract][Hide abstract] ABSTRACT: Gastric carcinogenesis is a multifactorial H.pylori-triggered dynamic process that goes through a cascade of preneoplastic conditions. The expression of miRNAs in the stomach with regard to preneoplastic precursor conditions and H.pylori infection has not been investigated systematically. In this prospective proof-of-principle study, we evaluated the miRNA expression in gastric antrum and corpus mucosa from patients with chronic non-atrophic gastritis (CNAG), atrophic gastritis (AG), and GC compared to controls. Gastric normal mucosa shows a unique expression pattern for miR-21, miR-155 and miR-223, which is specific for different regions. In correlation with progression of Correa's cascade and H.pylori infection, we observed a gradual increase in miR-155 and miR-223 both in corpus and antrum and miR-21 only in the antrum mucosa. Using miRNA expression we calculated a score that allowed us to discriminate patients with AG from subjects with normal mucosa with high diagnostic accuracy in testing and validation cohorts reproducibly. In summary, the expression pattern of miRNAs in the gastric mucosa is gradually increased with progression of Correa's cascade and H.pylori infection, suggesting miRNAs as potential biomarkers for preneoplastic precursor conditions. However, differences of miRNA expression between the gastric antrum and the corpus need to be considered in future studies. G astric cancer (GC) remains one of the leading health burdens accounting for up to 8.8% of all cancers and is the third leading cause of cancer-related deaths worldwide 1 . GC is a consequence of multiple factors including genetic predisposition, environmental factors, diet and aging. Among these, chronic inflam-mation due to H.pylori infection plays the key role in triggering carcinogenesis. According to a multistep process first described by Correa, development of intestinal type gastric cancer is triggered by H.pylori driven inflam-mation leading to typical stages of mucosal alterations such as chronic gastritis, glandular atrophy, intestinal metaplasia, dysplasia before reaching the final stage of invasive GC 2
[Show abstract][Hide abstract] ABSTRACT: To assess whether antibiotic resistance varies between the antrum and corpus of the stomach of patients that are either Helicobacter pylori (H. pylori) therapy-naive or pre-treated.
H. pylori strains were isolated from antrum and corpus biopsies from 66 patients that received a diagnostic gastroduodenoscopy for variant clinical indications. Antimicrobial susceptibility to amoxicillin, clarithromycin, tetracycline, metronidazole, levofloxacin and rifabutin was tested with the E-test method on Iso-Sensitest agar with 10 vol% defibrinated horse blood. In patients with a different antibiotic susceptibility pattern between the isolates from the antrum and corpus, DNA fingerprinting via random amplified polymorphic DNA analysis was performed to detect differences among DNA patterns of H. pylori isolates.
Primary, secondary and tertiary resistance to clarithromycin was 6.9%, 53.8% and 83.3%, retrospectively. Metronidazole and levofloxacin resistance also increased according to the number of previous treatments (17.2%, 69.2%, 83.3%; 13.8%, 23.1%, 33.3%). Tertiary resistance to rifabutin was detected in 12.5% of patients. In none of the 66 patients a resistance against amoxicillin or tetracycline was detectable. Discordant antibiotic susceptibility between antrum and corpus isolates for different antibiotics was seen in 15.2% (10/66) of the patients. Two out of those ten patients were naive to any H. pylori antibiotic treatment. The remaining eight patients previously received at least one eradication therapy. DNA fingerprinting analysis revealed no substantial differences among DNA patterns between antrum and corpus isolates in the majority of patients suggesting an infection with a single H. pylori strain.
Different antibiotic susceptibility between antrum and corpus biopsies is a common phenomenon and a possible explanation for treatment failure. Resistant H. pylori strains may be missed if just one biopsy from one anatomic site of the stomach is taken for H. pylori susceptibility testing.
World journal of gastroenterology : WJG. 11/2014; 20(43):16245-51.
[Show abstract][Hide abstract] ABSTRACT: Cathepsin K (CatK) is mainly expressed by osteoclasts and plays an important role in bone resorption. As CatK is expressed and secreted by osteoclasts during active bone resorption, it may be a useful and specific biochemical marker of osteoclastic activity. Therefore, CatK serum levels were studied for monitoring the treatment of females with postmenopausal osteoporosis by zoledronic acid. The serum CatK levels were determined in nine postmenopausal females before and after 3, 6 and 12 months of treatment. The levels were significantly reduced after 3 and 6 months (P<0.05), whereas they returned to baseline after 1 year. Taken together, the serum level of CatK may be suitable for monitoring anti-osteoporotic therapy in association with treatment response.
[Show abstract][Hide abstract] ABSTRACT: Intracellular zinc homeostasis is tightly regulated under physiological conditions; however, dysregulation of zinc levels has been reported in various chronic inflammatory and malignant diseases. In this study, we aimed to assess the expression pattern of the 24 currently known zinc transporters in resting and stimulated human peripheral blood mononuclear cells (PBMCs). The cells were isolated from healthy probands and subsequently stimulated with phytohaemagglutinin (PHA) for 3 days. The expression levels of zinc transporters [Zrt/IRT-like protein (ZIP) and cation diffusion facilitator/zinc transporter protein (CDF/ZnT) families] were analyzed by quantitative reverse transcription-polymerase chain reaction. Of the 24 genes encoding for zinc transporters, 19 were found to be ubiquitously expressed in PBMCs. ZIP5 and ZnT10 were not found in all 5 samples, whereas ZIP12, ZnT3 and ZIP2 were expressed in only 1-2 out of 5 PBMC samples. Of note, stimulation by PHA led to an overall downregulation of zinc transporters in the PBMCs of 4 out of the 5 subjects. Notably, the transcript levels of ZIP14 were consistently induced and those of ZIP3 and ZIP4 consistently downregulated in all 5 subjects, whereas the corresponding levels of the remaining 21 genes varied. Data from this study may facilitate a better understanding of the pathophysiological role of deregulated zinc transporters in chronic inflammatory diseases.
[Show abstract][Hide abstract] ABSTRACT: MicroRNAs (miRNAs) are known for their function as translational regulators of tumor suppressor or oncogenes. Single nucleotide polymorphisms (SNPs) in miRNAs related genes have been shown to affect the regulatory capacity of miRNAs and were linked with gastric cancer (GC) and premalignant gastric conditions. The purpose of this study was to evaluate potential associations between miRNA-related gene polymorphisms (miR-27a, miR-146a, miR-196a-2, miR-492 and miR-608) and the presence of GC or high risk atrophic gastritis (HRAG) in European population.
Gene polymorphisms were analyzed in 995 subjects (controls: n = 351; GC: n = 363; HRAG: n = 281) of European descent. MiR-27a T>C (rs895819), miR-146a G>C (rs2910164), miR-196a-2 C>T (rs11614913), miR-492 G>C (rs2289030) and miR-608 C>G (rs4919510) SNPs were genotyped by RT-PCR.
Overall, SNPs of miRNAs were not associated with the presence of GC or HRAG. We observed a tendency for miR-196a-2 CT genotype to be associated with higher risk of GC when compared to CC genotype, however, the difference did not reach the adjusted P-value (odds ratio (OR) - 1.46, 95% confidence interval (CI) 1.03-2.07, P = 0.032). MiR-608 GG genotype was more frequent in GC when compared to controls (OR -2.34, 95% CI 1.08-5.04), but significance remained marginal (P = 0.029). A similar tendency was observed in a recessive model for miR-608, where CC + CG vs GG genotype comparison showed a tendency for increased risk of GC with OR of 2.44 (95% CI 1.14-5.22, P = 0.021). The genotypes and alleles of miR-27a, miR-146a, miR-196a-2, miR-492 and miR-608 SNPs had similar distribution between histological subtypes of GC and were not linked with the presence of diffuse or intestinal-type GC.
Gene polymorphisms of miR-27a, miR-146a, miR-196a-2, miR-492, miR-492a and miR-608 were not associated with the presence of HRAG, GC or different histological subtypes of GC in European subjects.
PLoS ONE 01/2014; 9(1):e87467. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Different strategies of colonization or infection by E. coli result in formation of certain adhesion patterns which help also in classifying intestinal E. coli into pathotypes. Little is known about adhesion patterns and host- and tissue adaption of commensal E. coli and about E. coli originating in clinically healthy hosts carrying pathotype-specific virulence-associated genes.
Adhesion pattern of E. coli (n = 282) from humans and from 18 animal species were verified on intestinal human Caco-2 and porcine IPEC-J2 cells and, furthermore, for comparison on human urinary bladder 5637, porcine kidney PK-15 epithelial and HEp-2 cells. The analysis was carried out on 150,000 images of adhesion assays.Adhesion patterns were very diverse; 88 isolates were completely non-adherent, whereas 194 adhered to at least one cell line with the dominant adhesion patterns "diffusely distributed" and "microcolony formation". Adhesion patterns "chains" and "clumps" were also visible. Chain formation was mediated by the presence of epithelial cells. Clump formation was very specific on only the 5637 cell line. All enteropathogenic (eae+) E. coli (EPEC; n = 14) were able to form microcolonies which was cell line specific for each isolate. Most EPEC formed microcolonies on intestinal IPEC-J2 and Caco-2 but several also on urinary tract cells. Shigatoxin-producing (stx+) E. coli (n = 10) showed no specific adhesion patterns.
E. coli isolates were highly diverse. Commensal and pathogenic isolates can adhere in various forms, including diffuse distribution, microcolonies, chains and clumps. Microcolony formation seems to be a global adhesion strategy also for commensal E. coli.
Gut Pathogens 11/2013; 5(1):31. · 2.07 Impact Factor
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[Show abstract][Hide abstract] ABSTRACT: Recently, there has been a growing interest in an expanding group of cytokines known as "IL-12 family". The so far gained knowledge about these cytokines, as crucial playmakers in mucosal immunity, has not yet been sufficiently investigated in the context of Helicobacter pylori infection. All genes encoding the monomeric components of these cytokines and their corresponding receptors were examined in gastric epithelial cell lines (AGS and MKN-28) after being infected with 4 H. pylori strains: BCM-300, P1 wild-type, and P1-derived isogenic mutants lacking cytotoxin-associated gene A (cagA) or virulence gene virB7 (multiplicity of infection=50). Both infected and uninfected samples were analyzed after 24h and 48h using real-time quantitative polymerase chain reaction (RT-qPCR). Gene expression analysis demonstrated a strong upregulation of IL23A (encodes p19) by infection, whereas IL23R, Epstein-Barr virus-induced gene 3 (EBI3), IL6ST, IL12A, and IL27RA were found to be expressed, but not regulated, or to a lesser extent. Transcripts of IL12RB2, IL12B, IL12RB1, and IL27A were not detected. Interestingly, P1 resulted in stronger alterations of expression than CagA mutant and BCM-300, particularly for IL23A (59.7-fold versus 32.4- and 6.7-fold, respectively in AGS after 48h, P<.05), whereas no changes were seen with VirB7 mutant. In a proof-of-principle experiment, we demonstrated epithelial-derived expression of IL-12, p19, and Ebi3 in gastric mucosa of gastritis patients using immunohistochemistry (IHC). Unlike IL-12 and Ebi3, increased immunostaining of p19 was observed in H. pylori gastritis. Herein, we highlight the potential role of gastric epithelial cells in mucosal immunity, not only because they are predominant cell type in mucosa and initial site of host-bacterial interaction, but also as a major contributor to molecules that are thought to be primarily expressed by immune cells so far. Of these molecules, p19 was the most relevant one to H. pylori infection in terms of expression and localization.
PLoS ONE 09/2013; 8(9):e75192. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Proton pump inhibitor (PPI)-refractory heartburn may be due to persistent gastro-oesophageal reflux, oesophageal hypersensitivity or functional heartburn (FH). The differentiation between non-erosive reflux disease (NERD) and FH may be very difficult. However, this differentiation is important for appropriate therapeutic management. Dilated intercellular spaces (DIS), papillary elongation (PE) and basal cell hyperplasia (BCH) can be all assessed by light microscopy. Whether these mucosal abnormalities allow the differentiation of NERD from FH in PPI-refractory patients is uncertain.
To assess histopathological findings by light microscopy in patients with refractory heartburn to differentiate NERD from FH.
Sixty-two patients with PPI-refractory symptoms underwent EGD and MII-pH after pausing PPI medication for 2 weeks before investigation. Twenty-five subjects without upper gastrointestinal symptoms were included as controls. Symptom assessment was based on the reflux disease questionnaire (RDQ). Biopsies were taken 3-5 cm above the gastro-oesophageal junction. DIS, PE, BCH and infiltration of immune cells were evaluated and a sum score was calculated.
Based on endoscopy and MII-pH, GERD was diagnosed in 43 patients (NERD: 20; ERD: 23) and FH in 19 patients. There was no difference in symptoms between the groups. Each individual histopathological item was different between the groups (P < 0.0001). Between NERD and FH, the most significant difference was found for DIS and the histopathological sum score (P < 0.001).
These findings suggest that oesophageal biopsies are useful to differentiate NERD from FH. Increased DIS and a histological sum score are the most significant histopathological abnormalities in NERD as compared with FH.
[Show abstract][Hide abstract] ABSTRACT: Intestinal colonization is influenced by the ability of the bacterium to inhabit a niche which is based on the expression of colonization factors. E. coli carries a broad range of virulence-associated genes (VAGs) which contribute to intestinal (inVAGs) and extraintestinal infection (exVAGs). Moreover, initial evidence indicates that inVAGs and exVAGs support intestinal colonization.We developed new screening tools to genotypically and phenotypically characterize E. coli originating in humans, domestic pigs and 17 wild mammalian and avian species. We analyzed 317 isolates for the occurrence of 44 VAGs using a novel multiplex PCR microbead assay (MPMA), and for adhesion to four epithelial cell lines using a new adhesion assay. We correlated data for the definition of new adhesion genes.InVAGs were identified only sporadically, particularly in roe deer (Capreolus capreolus) and European hedgehog (Erinaceus europaeus). The prevalence of exVAGs depended on isolation from a specific host. Human UPEC carried exVAGs with the highest prevalence followed by badger (Meles meles) and roe deer isolates. Adhesion was found to be very diverse. Adhesion was specific to cells, host and tissue, though also unspecific. Occurrence of the following VAGs was associated with a higher adhesion rate to one or more cell lines: afa/dra/daaD, tsh, vat, ibeA, fyuA, mat, sfa/foc, malX, pic, irp2 and papC.Summarized, we established new screening methods which enabled us to characterize large amounts of E. coli. We defined reservoirs for potential pathogenic E. coli. We also identified a very broad range of colonization strategies and defined potential new adhesion genes.
Applied and Environmental Microbiology 07/2013; · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Helicobacter pylori eradication rates show a constant decline over the last few years. The main reason for H. pylori treatment failure is the increasing antibiotic resistance.We assessed antibiotic susceptibility of H. pylori in a region of mid-Germany and analyzed the relationship of antibiotic resistance with the number of eradication therapies over a period of 7 years (2005-2012).
H. pylori strains were isolated from 436 patients who underwent gastroscopy for different clinical indications. Susceptibility to amoxicillin, clarithromycin, metronidazole, tetracycline, levofloxacin, and rifabutin was determined using the E-test.
Primary, secondary, and tertiary resistances against clarithromycin were 7.5, 63.2, and 75.4%, respectively. Primary, secondary, and tertiary resistances to levofloxacin were 11.7, 17.6, and 36.4% and to metronidazole were 32.7, 63.2, and 80.1%, respectively. The resistance rates against tetracycline and rifabutin were comparatively low (<5%), even in patients with previous exposure to these antibiotics. Resistance to rifabutin increased to 6.2% in patients who received more than two previous eradication therapies. Amoxicillin resistance was not detectable in all patients.
In our region, we observed a stable, but constantly increasing, resistance rate to antibiotics commonly used for the treatment of H. pylori infection. Knowledge of the local antibiotic resistance rates is essential for developing successful treatment strategies for H. pylori eradication.
European journal of gastroenterology & hepatology 07/2013; · 1.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Experimental medicine has evolved tremendously in the last few years. In particular, the introduction of novel techniques, in-vitro models, knock-out/transgenic animals and high-through put analytical methodologies have resulted in a deeper understanding of cellular pathophysiology and diseases. The daily clinical management has benefited by the introduction of biomarkers and targeted therapies. This development has been accompanied by increasing specialisation across all fields of research and medicine. Therefore, clinical-translational research requires a team of competent partners nowadays. The visceral surgeon can contribute significantly to these projects. The present review highlights several aspects of translational research and put chances and potential pitfalls into perspective in context with the work of the visceral surgeon.
Zentralblatt für Chirurgie 05/2013; · 0.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The interplay between gastric and intestinal transcription factors has an important impact on gastric carcinogenesis. We compared the gene expression of CDX1, CDX2, SOX2 and related downstream genes in tumour and tumour surrounding gastric tissue of 48 gastric cancer patients with 30 healthy controls. There was no difference of gene expression of CDX1 and CDX2 between tumour or tumour-adjacent and tumour-distant mucosa, but both factors were significantly higher expressed in cancer patients compared with controls (p<0.01). SOX2 was downregulated in tumour tissue compared to controls, whereas tumour-adjacent and tumour-distant mucosa showed intermediate SOX2 expression. Laurén type and Helicobacter pylori infection had no significant impact on expression of the transcription factors. Expression of CDX1 and CDX2 was higher in the presence of intestinal metaplasia. The differential regulation of the gene expression of CDX1, CDX2 and SOX2 in patients with gastric cancer affects not only the tumour but also the non-neoplastic tumour-distant mucosa.
Journal of clinical pathology 04/2013; 66(9). · 2.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVES: C-telopeptide crosslaps (CTX) and bone-specific alkaline phosphatase (BAP) do not provide sufficient sensitivity and specificity for diagnosis of osteoporosis. Cathepsin K (CatK), osteoprotegerin (OPG), and receptor activator of nuclear factor κB ligand (total (t) and soluble (s) RANKL) play an important role in bone metabolism. Thus serum levels of biochemical markers, each or in combination, may be useful in diagnosis of osteoporosis. STUDY DESIGN: In total, 121 healthy women, 27 premenopausal women aged between 20 and 45 years, and 94 postmenopausal women aged 59-81 years, all free of known skeletal disorders were included. They underwent bone density measurement and measurement of biochemical markers. MAIN OUTCOME MEASURES: Based on WHO criteria, women were stratified in four groups (premenopausal: healthy; postmenopausal: healthy, osteopenia, osteoporosis), and their levels of CatK, OPG, RANKL, CTX and BAP were analyzed. RESULTS: Using WHO criteria 21 postmenopausal women had normal bone mineral density (BMD), 49 had osteopenia and 24 had osteoporosis. There were no significant correlations of CatK, OPG and RANKL with BMD (T-score) in age-adjusted analysis, but for BAP and CTX. ROC analyses resulted in poor diagnostic validity of all parameters. The best result - also confirmed by discriminant analysis - was yielded by BAP (AUC=0.646 [0.510; 0.781]). A combination of variables did not significantly improve the diagnostic power. CONCLUSIONS: Baseline serum levels of BAP, CTX, CatK, OPG, sRANKL or tRANKL alone or in combination are not suitable to distinguish osteoporotic from non-osteoporotic postmenopausal women with sufficient accuracy.
[Show abstract][Hide abstract] ABSTRACT: AIM: Noninvasive methods for assessment and follow-up of
hepatic fibrosis are important for the management of patients with
chronic liver disease. Our aim was to assess a new panel of surrogate
biomarkers for prediction of severe hepatic fibrosis in patients with
chronic liver disease of different aetiology.
METHODS: 118 patients [62 males (52.5%) and 56 females]
(47.5%) were prospectively enrolled with a mean age of 55.6 years
±14.9. The aetiology of chronic liver disease was hepatitis B virus
infection (n=12), hepatitis C virus infection (n=20), autoimmune
hepatitis (n=36), alcoholic steatohepatitis (n=10), non-alcoholic
steatohepatitis: (n=12), hepatocellular carcinoma (n=16). 12 patients
had no evidence of liver disease. Biomarkers of hepatic fibrosis and
liver function tests (α2-macroglobulin, haptoglobin, apolipoprotein
A1, total bilirubin, GGT, ALT, total cholesterol, AST, albumin,
CA19-9, CA125, CA 15-3, INR, platelet count, hyaluronic acid, nitric
oxide) were analyzed in serum. As reference for staging of fibrosis
we used FibroTest and FibroScan. Biomarkers were correlated to
hepatic fibrosis by univariate and multivariate analyses as well as
RESULTS: Univariate and multivariate analysis indicated that
platelet count, α2-macroglobulin, total bilirubin, GGT and total
cholesterol were the most relevant biomarkers related to the stage of
hepatic fibrosis. A new panel for prediction of severe hepatic fibrosis
was created using these relevant parameters. Applying this panel;
severe hepatic fibrosis was predicted with a sensitivity of 97.4%
and a specificity of 85.9% in comparison with FibroTest. Also a
sensitivity of 78.8% and specificity of 90.9% was obtained by the
panel in comparison to FibroScan.
CONCLUSION: The new noninvasive panel allows accurate
prediction of severe liver fibrosis in different types of chronic liver
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Gastroesophageal reflux disease (GERD) is associated with impaired epithelial barrier function that is regulated by cell-cell contacts. The aim of the study was to investigate the expression pattern of selected components involved in the formation of tight junctions in relation to GERD. METHODS: Eighty-four patients with GERD-related symptoms with endoscopic signs (erosive: n = 47) or without them (non-erosive: n = 37) as well as 26 patients lacking GERD-specific symptoms as controls were included. Endoscopic and histological characterization of esophagitis was performed according to the Los Angeles and adapted Ismeil-Beigi criteria, respectively. Mucosal biopsies from distal esophagus were taken for analysis by histopathology, immunohistochemistry and quantitative reverse-transcription polymerase chain reaction (RT-PCR) of five genes encoding tight junction components [Occludin, Claudin-1, -2, Zona occludens (ZO-1, -2)]. RESULTS: Histopathology confirmed GERD-specific alterations as dilated intercellular spaces in the esophageal mucosa of patients with GERD compared to controls (P < 0.05). Claudin-1 and -2 were 2- to 6-fold upregulation on transcript (P < 0.01) and in part on protein level (P < 0.015) in GERD, while subgroup analysis of revealed this upregulation for ERD only. In both erosive and non-erosive reflux disease, expression levels of Occludin and ZO-1,-2 were not significantly affected. Notably, the induced expression of both claudins did not correlate with histopathological parameters (basal cell hyperplasia, dilated intercellular spaces) in patients with GERD. CONCLUSIONS: Taken together, the missing correlation between the expression of tight junction-related components and histomorphological GERD-specific alterations does not support a major role of the five proteins studied in the pathogenesis of GERD.