Asia-Pacific consensus guidelines on gastric cancer prevention.

BACKGROUND AND AIM: Gastric cancer is a major health burden in the Asia-Pacific region but consensus on prevention strategies has been lacking. We aimed to critically evaluate strategies for preventing gastric cancer. METHODS: A multidisciplinary group developed consensus statements using a Delphi approach. Relevant data were presented, and the quality of evidence, strength of recommendation, and level of consensus were graded. RESULTS: Helicobacter pylori infection is a necessary but not sufficient causal factor for non-cardia gastric adenocarcinoma. A high intake of salt is strongly associated with gastric cancer. Fresh fruits and vegetables are protective but the use of vitamins and other dietary supplements does not prevent gastric cancer. Host-bacterial interaction in H. pylori infection results in different patterns of gastritis and differences in gastric acid secretion which determine disease outcome. A positive family history of gastric cancer is an important risk factor. Low serum pepsinogens reflect gastric atrophy and may be useful as a marker to identify populations at high risk for gastric cancer. H. pylori screening and treatment is a recommended gastric cancer risk reduction strategy in high-risk populations. H. pylori screening and treatment is most effective before atrophic gastritis has developed. It does not exclude the existing practice of gastric cancer surveillance in high-risk populations. In populations at low risk for gastric cancer, H. pylori screening is not recommended. First-line treatment of H. pylori infection should be in accordance with national treatment guidelines. CONCLUSION: A strategy of H. pylori screening and eradication in high-risk populations will probably reduce gastric cancer incidence, and based on current evidence is recommended by consensus.

Pepsin like activity in bronchoalveolar lavage fluid is suggestive of gastric aspiration in lung allografts.

BACKGROUND: A biologically plausible link between gastro-oesophageal reflux (GOR), aspiration, and lung allograft dysfunction has been suggested, but there is no systematic evidence indicating the presence of gastric contents in the lung. We have tested the hypothesis that pepsin, as a marker of aspiration, is detectable in bronchoalveolar lavage (BAL) fluid of allograft recipients who had not reported symptoms of GOR. METHODS: Standardised 3 x 60 ml surveillance BAL fluid samples from 13 chronologically sequential stable lung allograft recipients without chronic rejection (10 patients treated with a prophylactic proton pump inhibitor) were studied. Lavage supernatants were assayed by an ELISA based on a monospecific goat antibody for pepsin/pepsinogen. Pepsin levels were compared with those from four normal volunteer controls. RESULTS: Pepsin levels were measurable in all allograft recipients, in keeping with gastric aspiration (median 109 ng/ml (range 35-1375)). In the control group the pepsin levels were below the limit of detection. Treatment with a proton pump inhibitor was not correlated with pepsin levels. There was no correlation between BAL fluid neutrophils and pepsin levels. CONCLUSION: These data demonstrate lung epithelial lining fluid concentrations of pepsin in lung allograft recipients which are much higher than blood reference levels, with no detectable pepsin in controls. This provides direct evidence of gastric aspiration, which is potentially injurious to the allograft.

Electrotitration curves of human gastric pepsinogens in agarose gels.

Electrotitration curves (ETC) of a marker protein mixture, pH 2.5-5.65, and human pepsinogens were performed in an agarose gel, containing 2% acid carrier ampholytes, forming a pH range of 2.5-5. Although the establishment of the pH gradient by isoelectric focusing was not quite complete and linear, both biochemically and immunochemically different types of pepsinogen C (PGC) and pepsinogen A (PGA) zymogens as well as the acid isoelectric points (pI) marker proteins were separated with good resolution. Three main fractions of PGA (Pg3, Pg4, and Pg5) were detected. To obtain an exact determination of the pepsinogen pIs, a simple and very fast 10 s pressure blot technique was applied. Human pepsinogens were separated alone or mixed with pI marker proteins in the pH range 2.4-5.65. No effect of the markers was observed on the pepsinogen migration. To visualize the different protein samples in the gel and on nitrocellulose membrane, we have used colloidal gold (AuroDye) staining, proteolytic activity, and immunostaining with monoclonal antibodies anti PGA and PGC. The described method shows an ability to separate proteins at acidic conditions with a resolution comparable to isoelectric focusing with immobilized pH gradients, but much faster, easier, and cheaper. In addition, the technique allows us to determine precise and exact pI values, and is suitable for studies of the pepsinogen polymorphism and its role in gastric diseases.

Polyclonal antibodies for immunochemical determination of human pepsinogens.

The human gastric mucosa contains two main groups of aspartic proteinases, pepsin (EC 3.4.23.1) and gastricsin (EC 3.4.23.3), which differ by their structural, kinetic and immunological characteristics. The ratios between human aspartic proteases are important from the diagnostic point of view. Rabbit polyclonal antisera against human pepsinogen A and pepsinogen C (progastricsin) were obtained and tested for clinical purposes. Immunoblotting procedure seems to be a simple and sufficiently sensitive method for qualitative determination of pepsinogens in human gastric mucosa.

Establishment and characterization of two cell lines from N-methyl-N-nitrosourea-induced mouse glandular stomach carcinomas.

We previously reported the induction with N-methyl-N-nitrosourea (MNU) of mouse glandular stomach carcinomas showing a gastric phenotype but variation in histologic appearance, as with human gastric carcinomas. In the present study, we established two cell lines, designated MGT-40 and MGT-93, from MNU-induced mouse glandular stomach carcinomas. These cell lines are keratin-positive and grow as epithelial monolayers in culture, requiring transforming growth factor alpha, epidermal growth factor or insulin/transferrin for optimal growth in addition to 10% fetal bovine serum. Retention of the differentiated phenotype for gastric surface mucous cells has been confirmed by cathepsin E immunohistochemistry and reverse transcriptase-polymerase chain reaction for mouse spasmolytic polypeptide. Neither transplantability in nude mice nor colony formation on soft agar was observed, except in one subline. Chromosome analysis revealed aneuploidy with modal chromosome numbers ranging from 58 to 78 and no specific structural abnormalities. This is the first report of cell lines derived from mouse glandular stomach carcinomas. They should prove useful for studies of the mechanisms of regulation of growth and differentiation.

Changes in the zymogenic cell populations of the abomasa of sheep infected with Haemonchus contortus.

The effects of dietary urea supplementation and of a 10-week trickle infection regime, simulating chronic exposure to Haemonchus contortus, on the zymogenic population of the abomasa of Hampshire Down lambs was examined. At necropsy a variety of parameters including plasma pepsinogen concentration, the wet weights of abomasal fundic mucosal pieces and the amounts of pepsinogen contained in them, were assessed. Tissue pepsinogen concentration was measured as the total, acid-stable proteolytic activity present in mucosal homogenates, as well as immunohistochemically. The immunohistochemical findings were quantified using computer-aided image analysis. Elevation of plasma pepsinogen concentrations in infected animals was of borderline significance (P = 0.06). The fundic mucosae of infected animals were heavier (P < 0.02) than those of control animals, but there was no overall change in the pepsinogen content of tissues. Immunohistochemistry revealed that infected animals had increased numbers of zymogenic cells, due to mucous cell hyperplasia and the adaptation of cells to produce both mucins and pepsinogen. The pepsinogen content of chief cells, the major source of pepsinogen in uninfected animals, was reduced in infected lambs. Image analysis confirmed that at a mid-point of the mucosa of infected animals there was increased pepsinogen-specific immunoreactivity that corresponded with areas of mucosal hyperplasia. Mucous cell hyperplasia might therefore allow the maintenance of pepsinogen secretion in infected animals even if chief cell output is reduced.

Effects of Helicobacter pylori eradication on gastric function indices in functional dyspepsia. A prospective controlled study.

BACKGROUND: To date, it is unclear whether Helicobacter pylori infection is associated with disturbances of gastric emptying or acid secretion in patients with functional dyspepsia (FD). Our aim was to investigate whether, in the long run, cure of H. pylori infection significantly influences gastric emptying of solids, acid secretion, and gastrin and pepsinogen I (PGI) release in patients with FD. METHODS: Thirty-eight consecutive H. pylori-positive patients with FD, whose complaints were scored for severity and frequency on the basis of a validated symptom questionnaire, were initially enrolled in the study. They were randomized to receive an eradicating regimen consisting of omeprazole plus clarithromycin and tinidazole for 1 week or full-dose ranitidine for 3 weeks. In 33 patients (18 H. pylori-cured and 15 with persistent infection) basal and pentagastrin-stimulated acid secretion, fasting and meal-induced gastrin concentrations, fasting serum PGI levels, and gastric emptying of solids were determined before and 6 months after therapy. RESULTS: In the 18 H. pylori-cured patients meal-induced gastrin and fasting PGI levels were significantly reduced after 6 months as compared with pretreatment values (peak serum gastrin, 76.0 +/- 23.4 versus 111.9+/-37.4 pg/ml; PGI, 57.1+/-23.4 versus 72.9+/-29.1 ng/ml), whereas they remained virtually unchanged in the 15 patients with persistent infection. In contrast, both basal and stimulated acid secretion and gastric emptying time of solids remained unmodified over time in both groups of patients. CONCLUSIONS: We confirm that also in patients with functional dyspepsia H. pylori eradication in the long run significantly reduces gastrin and PGI release as a result of improvement in the underlying antral gastritis, but this is not accompanied by modifications of gastric emptying of solids or acid secretion.

Immunocytochemistry and in situ hybridization studies of pepsinogen C-producing cells in developing rat fundic glands.

The ontogeny of pepsinogen C-producing cells in rat fundic glands was studied by means of light and electron microscopy using an antiserum raised against a synthetic peptide based on rat pepsinogen C. To confirm the immunocytochemistry results, the expression of rat pepsinogen C messenger RNA (mRNA) in the fundic gland was also examined by in situ hybridization using a digoxigenin-labeled RNA probe. In adult rats, pepsinogen C was produced by chief cells, mucous neck cells, and intermediate mucopeptic cells. Pepsinogen C-producing cells appeared in embryos as early as 18.5 days' gestation. The development of these cells could be classified into four stages: (1) 18.5 days' gestation to 0.5 days after birth; (2) 0.5 days to 2 weeks after birth; (3) 3-4 weeks after birth; (4) 4-8 weeks after birth. In embryos and young animals, pepsinogen C-producing cells were mucopeptic cells. By 4 weeks after birth, mucous neck cells could be distinguished morphologically. The maturation stages of the chief cells could be traced by electron microscopy along the longitudinal axis of the rat fundic gland by double-staining with anti-pepsinogen C antibody and periodic acid-thiocarbohydrazide-silver proteinate. Positive reactions for pepsinogen C and pepsinogen C mRNA expression were detected in mucous neck cells. Therefore, we conclude that mucous neck cells are precursor cells of chief cells. Mucous neck cells, intermediate cells, and chief cells are in the same differentiating cell lineage.

A new view of the so-called adenoma malignum of the uterine cervix.

Adenoma malignum of the uterine cervix (mucinous type of minimal deviation adenocarcinoma, mucinous MDA), is a unique neoplasm that is difficult to diagnose owing to the deceptively benign appearance of the tumour cells. The present study was undertaken to explore the phenotypic expression of this tumour compared with those of non-neoplastic cervical tissues and of cervical carcinomas of various types. Ten cases of mucinous MDA, 50 cases with non-neoplastic cervical tissues, 13 of cervical adenocarcinoma including the mucinous (endocervical or intestinal type) and endometrioid types, and 2 of mucoepidermoid carcinoma were examined by various histochemical staining methods, including those for gastric mucins, pepsinogen, lysozyme, chromogranin A and carcinoembryonic antigen. The results revealed that mucinous MDA characteristically exhibited gastric phenotypes. The presence of gastric metaplasia was also demonstrated in 9 cases of mucinous MDA and in 5 of the other cases examined. The 7 endocervical-type adenocarcinomas also included 4 that expressed gastric phenotypes, and 2 of the 3 intestinal-type adenocarcinomas showed the same properties focally. These results indicate the presence of a group of lesions expressing gastric phenotypes in the uterine cervix and suggest a close relationship between these lesions. Cervical adenocarcinomas expressing gastric phenotypes are probably derived from MDA.

Expression of intrinsic factor and pepsinogen in the rat stomach identifies a subset of parietal cells.

Morphological and functional heterogeneity of parietal cells has been thought to be due to different maturation positions within the gastric gland. Morphodynamic studies have shown that 2% of parietal cells in mice derive from a pre-neck (chief) cell precursor. Intrinsic factor (IF) and pepsinogen, markers of rat chief cells, were used to determine if these proteins identified a subset of parietal cells that might reflect origin from the pre-neck cell lineage. The zymogenic region of the rat stomach and gradient-isolated fractions enriched in parietal and chief cells were fixed in 10% buffered Formalin or in Bouin's solution. Immunostaining was performed using indirect immunoperoxidase histochemistry and double-labeled immunofluorescence with antibodies raised against human IF, pepsinogen II, and H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase). In intact tissue, parietal (H(+)-K(+)-ATPase-positive) cells were found starting at the upper edge of the isthmus, but parietal cells positive for IF and pepsinogen were only found from just below the isthmus and neck region to the base of the gastric gland. Three to four percent of isolated parietal cells were positive for these ectopic markers. This subset of cells was also positive for H(+)-K(+)-ATPase. Thus products of rat chief cells are expressed in a subset of parietal cells. The percentage of positive cells is similar to that predicted to be derived from the pre-neck (chief) precursor lineage in the mouse. The distribution of these cells to the lower neck and base of the gland suggests that the expression of chief cell products is consistent with either predetermination by lineage or parietal cell maturation or with both processes.

Effect of chemoprophylaxis with an ivermectin sustained-release bolus on acquired resistance to gastrointestinal parasites in cattle.

The influence of chemoprophylaxis with an ivermectin sustained-release bolus in the first grazing season on the resistance of cattle to gastrointestinal nematodes during the following grazing season was investigated. In 1993 and 1994 dairy replacement calves were either given one bolus at the start of their first grazing season or left untreated. The two groups were grazed separately on a pasture that was divided into two similar sized paddocks. Faecal egg counts, serum pepsinogen and antibody levels were measured to evaluate host-parasite contact. Pasture infection levels were estimated by pasture larval counts and worm counts in tracer calves. After winter housing the animals were monitored during their second grazing season on a pasture that was also divided into two similar sized paddocks. Acquired resistance to gastrointestinal nematodes was evaluated by faecal egg counts and weight gains. Again, pasture infection levels were determined and pepsinogen and antibody levels were measured. During the first grazing seasons gastrointestinal nematode infections were controlled very effectively by the bolus, as shown by the greater weight gains, the negligible faecal egg counts and the low serum pepsinogen and antibody levels in the treated calves. In contrast, all parameters showed extensive parasite-host contact in the untreated animals. The efficient prophylaxis in the treated groups resulted in low levels of larval contamination on the paddocks grazed by the treated animals, compared to moderate infection levels at the end of both first grazing seasons on the paddocks grazed by the untreated animals. During the second grazing seasons (1994 and 1995) the faecal egg output was low in all groups. Although in the previously treated animals faecal egg counts were consistently higher, the differences were minimal, resulting in comparable levels of larval contamination on both paddocks. Serum pepsinogen and antibody levels were not significantly different between the groups and indicated a similar level of larval uptake on both paddocks. No negative effect of the previous chemoprophylaxis on the clinical condition and the weight gain of the second season grazing animals was observed.

Early specification of intestinal epithelium in the chicken embryo: a study on the localization and regulation of CdxA expression.

CdxA, a chicken homeobox-containing gene related to caudal in Drosophila, has been implicated in the regionalization of endoderm. It is reported here that, in the development of the chicken embryo, CdxA expression appears in the endoderm at day 1.5 of development as bilateral bands on either side of the splanchnopleure which later contribute to intestinal epithelium. The CdxA-expressing area extends medially and caudally as formation of the gut tube progresses. It is also shown that the rostral limit of CdxA expression demarcates the boundary between stomach and duodenum after day 3 of development. CdxA is not expressed in digestive tract appendages which open into the intestine, such as pancreas, liver and allantois. Early restriction of CdxA expression in intestinal lineage suggests that the intestinal specification involving CdxA expression commences before the gut tube is formed. The expression of CdxA in epithelial-mesenchymal tissue recombinants suggests that mesenchymal influence regulating CdxA expression plays an important role in confirming the boundary between the stomach and intestine. Chronological change in the spatial distribution of CdxA transcripts and the results of tissue recombination experiments, together with precise fate maps of early endoderm and splanchnic mesoderm, lead to a model of mechanisms by which intestinal specification is brought about.

Immunofluorometric assay of pepsinogen C and preliminary clinical applications.

We developed mouse monoclonal antibodies (Abs) against pepsinogen C with highly purified antigen isolated from gastric mucosa. The Abs were used to construct a two-site sandwich-type assay for pepsinogen C with time-resolved fluorometry as a detection technique. The assay has a detection limit of 0.1 microgram/L and is precise (within-run and day-to-day CVs < 11%). We used this assay to measure pepsinogen C in seminal plasma, breast cyst fluid, amniotic fluid, male and female serum, serum from patients with prostate cancer, urine, breast tumor cytosolic extracts, breast milk, and cerebrospinal fluid. Highest pepsinogen C concentrations were in seminal plasma, followed by breast cyst fluid and amniotic fluid. We found no correlation between prostate-specific antigen concentrations and concentrations of pepsinogen C in serum of prostate cancer patients, and concluded that this marker is not useful for either diagnosing or monitoring prostatic carcinoma. The availability of a highly sensitive, reliable, and convenient method for quantifying pepsinogen C will allow investigations into the possible diagnostic value of this analyte in various clinical conditions, including benign breast diseases, breast cancer, fertility, and pregnancy.

[Liquid chromatographic separation of pepsinogens].

In this work, we reported a method for the purification of pepsinogens from human gastric mucosa with high pressure gel filtration chromatography (HPLC) and medium pressure anion exchange chromatography (MPLC). First, the two fractions of Pg1 and mixture of PGI and PGII were separated from pepsinogens with HPLC on a Bio-Sil SEC-125 column within 50min. Then the mixture of fractions of PGI and PGII was further separated with MPLC on a Bio-Scale Q2 column within 30min.

Chronic atrophic gastritis and Helicobacter pylori infection in primary biliary cirrhosis: a cross-sectional study with matching.

BACKGROUND/AIMS: Primary biliary cirrhosis (PBC) is a chronic liver disease characterized by exocrine gland impairment. Up to now there have been no reports dealing with gastric mucosa involvement in this autoimmune condition, which is frequently associated with Sjögren's syndrome. The aim of this study was to investigate the morphologic, biochemical and immunological features of the gastric mucosa in PBC. METHODS: A cross-sectional study with matching was performed. Thirty-three PBC patients (30 F, 3 M, mean age 58 years; 17 with stage II-III, and 16 with stage IV disease) and 33 sex- and age-matched dyspeptic controls were included. Six biopsy specimens from the fundus (2), body (2) and antrum (2) were taken from all patients and controls. A serological assessment was performed for each subject, i.e. pepsinogen A (PGA), pepsinogen C (PGC), gastrin (G), and antibodies against Helicobacter pylori (anti-Hp IgG). RESULTS: Endoscopic gastritis was found in 22 PBC patients (66.6%). There was no difference between PBC patients and controls regarding the percentage of subjects with mild, moderate, severe or atrophic gastritis (AG). There was no difference in gastric mucosal involvement between PBC subjects with or without secondary Sjögren's syndrome. A discrepancy was observed in the data obtained with respect to Helicobacter pylori (H. pylori) infection. H. pylori colonization was significantly more frequent in controls than in PBC patients (79% vs 49%, p < 0.002), but anti-Hp IgG were detected in the same percentage in the two groups (90% vs 83% respectively). There was no difference between the two groups in the PGA, PGC, PGA/PGC ratio, or gastrin. Eight PBC patients had esophageal varices. CONCLUSIONS: PBC patients are not characterized by chronic atrophic gastritis. Even though they present chronic gastritis with the same prevalence as dyspeptic controls, and show signs of previous H. pylori infection as frequently as dyspeptic patients, they are actually much less frequently infected. The reasons for this observation are unclear.

Fast detection of phosphorylation of human pepsinogen A, human pepsinogen C and swine pepsinogen using a combination of reversed-phase high-performance liquid chromatography and capillary zone electrophoresis for peptide mapping.

A combination of reversed-phase high-performance liquid chromatography (RP-HPLC) and capillary zone electrophoresis (CZE) was used for characterization of alpha-chymotryptic digests of human pepsinogen A, human pepsinogen C (both isolated from stomach mucosa of patients suffering from gastric cancer), swine pepsinogen and their dephosphorylated forms. Combining RP-HPLC and CZE for peptide mapping allowed to detect phosphorylations in molecules of the above mentioned gastric zymogens. We have found one phosphate group in the molecule of human pepsinogen A and two phosphate groups in the molecule of human pepsinogen C. The investigated sample was obtained from stomach mucosa of a patient suffering from gastric cancer. An increased number of phosphate groups in molecules of human pepsinogen seems to be associated with gastric cancer. The developed method represent a suitable tool for studying relationships between specific phosphorylations of proteins and cancerogenesis or potentially could serve for early diagnosis of gastric cancer.

Ultrastructural immunogold localization of osteopontin in human gastric mucosa.

We performed ultrastructural immunogold localization of osteopontin in the mucosa of human stomach. This adhesive glycoprotein was present in mucous and chief cells of the epithelial layer and in macrophages in the lamina propria. Parietal and endocrine cells of the epithelial layer and mast cells and plasma cells in the lamina propria did not contain osteopontin, serving as internal negative controls. Subcellular localizations of osteopontin included secretory granules and synthetic organelles in mucous and chief cells and phagolysosomes in macrophages. Extracellular concentrations of osteopontin were present in the glycocalyx and in an electron-lucent band between epithelial surface cells and the gastric lumen. Paracellular edema between the epithelium of the same cells was devoid of osteopontin. Immunogold localization of pepsinogen II was done to identify cells with mixed granule populations and contents of multicompartmental secretory granules. These studies revealed mucous cell granules and chief cell granules, each containing compartmentalized storage products, which included osteopontin and mucigen in mucous cells and osteopontin and pepsinogen II in chief cells. Cytochemical controls for the immunogold localizations were negative. The subcellular distribution of osteopontin in human gastric mucosa suggests possible roles for this glycoprotein in barrier function, host defense, and/or secretion.

[Development of the gastric glands of cats (Felis silvestris catus)]. / Die Entwicklung der Magendrüsen der Katze (Felis silvestris catus).

There are five stages in the development of the cat's gastric glands: 1. During the stage of the indifferent epithelium from day 19 to day 24, the anlage of the stomach develops with all layers; 2. The stage of gland formation from day 24 to day 41 is the beginning of the gland buds. They develop in connection with endocrine cells on day 34 into primitive oxyntic and primitive mucous cells. The latter form the basis for all other cells, including the surface mucous cells; 3. During the stage of gland evagination from day 42 to 55, the anlagen are separated into primitive pits and tubules, while the cells continue to differentiate and the first intermediate cells are seen; 4. The stage of gland branching from day 56 to birth is characterized by the formation of additional glands at the bottom of the pits which change the ordinary anlagen into branched glands. During this stage, the cardiac glands are formed; 5. In the stage of gland maturation from birth to the 9th week, the peptic cells are formed and the glands start functioning. The oxyntic cells show carbonic-anhydrase activity and signs of acid secretion, and, between the weeks 4 and 8, the peptic cells contain pepsinogen, producing a negative reaction to PAS and a positive reaction to HID. Mucous cells and mucous neck cells produce PAS- and AB-positive mucin.

Determination of phosphoserine in human pepsinogens.

It has previously been assumed that, in contrast to porcine pepsinogen, the human pepsinogens are not phosphorylated. The present investigations show that phosphorylation does contribute to the electrophoretic heterogeneity of the human pepsinogens. A new chromatographic method for analysis of phosphoamino acid was developed. Quantitative determinations of phosphoserine were carried out after hydrolysis in 6 mol l-1 HCl (4 h, 110 degrees C). The recovery value of an authentic sample of phosphoserine, treated in parallel with the unknown samples, was used for calculations.

High performance liquid chromatography enables fast determination of ratios between individual proteolytic enzymes in human gastric mucosa.

The human gastric mucosa contains five isozymogens of pepsinogen (pepsinogens PGA1-PGA5), two isozymogens of gastricsinogen (gastricsinogens PGC6-PGC7) and zymogens of cathepsins. Ratios between some individual pepsins or pepsinogens are very important from a diagnostic point of view. The ratio of pepsinogen 3 to pepsinogen 5 is significant marker of gastric cancer and gastric ulcer. High-performance ion-exchange chromatography is an easy and fast method for determination of ratios between individual proteolytic zymogens in human gastric mucosa and thus could serve for additional diagnosis of gastric diseases.