Tritiated thymidine autoradiographic study on histogenesis and spreading of intestinal metaplasia in human stomach.

The non-diseased portions of the antral mucosa of patients suffering from gastric cancer or ulcer were biopsied. The biopsy specimens were then labelled with 3H-thymidine in vitro, and distribution of the labelled epithelial cells in the normal pyloric and in the intestinalized mucosa was studied with autoradiography, and modes of histogenesis and spreading of the intestinal metaplasia were studied, and kinetic characteristics of the intestinalized mucosa were discussed. In the normal pyloric mucosa, the labelled cells were confined to the isthmus region (the middle one-third level of the mucosa), indicating that the surface epithelial and the pyloric glandular cells are normally replaced from the isthmus region. On the other hand, a zone of the labelled cells was found at the lower one-third level in the intestinalized mucosa. The absorptive and the goblet cells in the intestinalized mucosa appear to be renewed by about 70 hours in a fashion similar to that of the small intestine. Microscopic and autoradiographic analysis of the antral mucosa in the course of intestinalization indicates that the intestinal metaplasia begins in the isthmus region of the pyloric glandular tubules of an intact mucosa unaffected by gross injury through transformation of the generative cells from a pyloric to an intestinal pattern. This permits the pyloric lining cells to be replaced with intestinal villous cells and also permits the generative cell zone of the intestinal tubules to shift from the isthmus to the base of the gland until the process is complete. The downward shift of the intestinal tubules occurs in a framework of one of the branched pyloric glands and other glands disappear, resulting in a change of mucosal architectures of the antrum from a branched to a simple tubular gland. The intestinal metaplasia spreads in the mucosa through multi-focal (and sporadical) transformation of the neck generative cells in individual glandular tubules.     

Local and systemic immune responses in murine Helicobacter felis active chronic gastritis.

Helicobacter felis inoculated per os into germfree mice and their conventional non-germfree counterparts caused a persistent chronic gastritis of approximately 1 year in duration. Mononuclear leukocytes were the predominant inflammatory cell throughout the study, although polymorphonuclear cell infiltrates were detected as well. Immunohistochemical analyses of gastric mucosa from H. felis-infected mice revealed the presence of mucosal B220+ cells coalescing into lymphoid follicles surrounded by aggregates of Thy-1.2+ T cells; CD4+, CD5+, and alpha beta T cells predominated in organized gastric mucosal and submucosal lymphoid tissue, and CD11b+ cells occurred frequently in the mucosa. Follicular B cells comprised immunoglobulin M+ (IgM+) and IgA+ cells. Numerous IgA-producing B cells were present in the gastric glands, the lamina propria, and gastric epithelium. Infected animals developed anti-H. felis serum IgM antibody responses up to 8 weeks postinfection and significant levels of IgG anti-H. felis antibody in serum, which remained elevated throughout the 50-week course of the study.     

IN SITU ANALYSIS OF TISSUE DYNAMICS AND p53 EXPRESSION IN HUMAN GASTRIC MUCOSA

In situ tissue dynamics were studied in 12 cases of human gastric mucosa, including normal gastric body mucosa and gastric glands with intestinal metaplasia, obtained from gastrectomy specimens of adenocarcinoma. Cell proliferation was determined by Ki67 immunoreactivity. DNA fragmentation was studied in situ by TdT-mediated dUTP-biotin nick end labelling (TUNEL). In addition, p53 expression was examined by both immunohistochemistry and mRNA in situ hybridization. In the oxyntic gastric glands, Ki67 immunoreactivity was observed exclusively in the proliferative zone and TUNEL-positive cells were present predominantly in the surface foveolar epithelium. In the gastric glands with complete intestinal metaplasia, Ki67-positive cells were present in the lower portion of the glands and TUNEL-positive cells in the superficial epithelium. In the gastric glands with incomplete intestinal metaplasia, TUNEL-positive cells were detected in the lower gastric glands adjacent to cells immunoreactive for Ki67; the proportion of these gastric glands with TUNEL-positive cells (40 out of 108 glands) was significantly higher than for oxyntic glands (94 out of 620 glands) or for glands with complete metaplasia (31 out of 254 glands). Relatively strong p53 immunoreactivity and mRNA hybridization were also observed in the proliferative and apoptotic areas of gastric glands with incomplete intestinal metaplasia. These results indicate that incomplete intestinal metaplasia is associated with increased cell turnover and p53 overexpression, possibly in response to various noxious or DNA-damaging stimuli.     

Oral tonsils: an immunoperoxidase study

Tissue morphometry and the distribution of cells containing immunoglobulin (Ig), J chain and secretory component (SC) were studied in eight oral tonsils. Quantitative immunohistochemistry revealed that IgG-containing cells predominated in all lymphoid compartments (follicles, extrafollicular areas and reticular epithelium) and that the IgG:IgA:IgM class ratios of the overall tonsillar immunocyte population were 13:8:2. Cells containing IgD and IgE were rare. IgG immunocytes showed no significant localisation within a given lymphoid compartment, whereas IgA cells were found predominantly in extrafollicular areas, especially adjacent to surface epithelium, and IgM cells were in follicles. J chain was present within IgM and some IgA cells. Tonsillar crypt and surface epithelium was negative for SC, suggesting that these structures are not directly involved in local mucosal immunity.     

Glicentin-containing cells in intestinal metaplasia,adenoma and carcinoma of the stomach

Summary Glicentin-containing cells (Glic. cells) in intestinal metaplasia, adenoma and carcinoma of the stomach were examined using immunohistochemical techniques. Glic. cells first occurred in the gastric mucosa of the transitional area between metaplastic and intact gastric glands. They frequently showed hyperplasia or micronoduli in the budding area of the deeper metaplastic glands, but in completely intestinalized mucosa these endocrine cells decreased remarkably. Gastric adenomas with mild dysplasia had a good number of glicentin-immunoreactive cells which were located in the deeper adenoma glands. Gastrin- and somatostatin-positive cells were also detected in the adenomas. The incidence of glicentin-positive tumor cells was significantly higher in well differentiated adenocarcinoma than in poorly differentiated adenocarcinoma. Among the seven cases of scirrhous argyrophil cell carcinoma, three showed glicentin- and glucagon-immunoreactivity in the same area of the tumor. These findings suggest that the selective increase of Glic. cells in intestinal metaplasia may be closely related to the development of gastric adenoma. Glicentin positive tumor cells in gastric carcinomas can be regarded to be an expression of intestinal or fetal markers.     

肠上皮生化,异型增生及胃癌的细胞动力学研究

48 specimens of gastric mucosa, including those of normal mucosa, intestinal metaplasia, atypical hyperplasia and carcinoma, were studied with in vitro 3H-TdR double labeling autoradiographic technique. On the basis of cell kinetics, intestinal metaplasia might be divided into two types: Type A consisted of those cases in which the values of LI, Ts and Tc were approximately closer to those of normal mucosa. The labeling cells appeared in the lower two thirds of the intestinalized glands, including small intestinal type and the complete type of colonic intestinal metaplasia. In type B, the average values of LI were higher. Ts and Tc values approximated those found in carcinoma a. Type B consisted mostly of the incomplete type of colonic intestinal metaplasia. The above evidence suggests that type B intestinal metaplasia is precancerous.     

Distribution of IgA 1 and IgA 2 plasma cells in various normal human tissues and in the jejunum of plasma IgA-deficient patients.

The distribution of IgA 1 and IgA 2 plasma cells was studied in normal human tissues. IgA 2 is a minor constituent in peripheral lymph nodes as well as in serum and in bone marrow plasma cells. An increased proportion of IgA 2 plasma cells was observed in gastric and intestinal mucosa, as well as in bronchial mucosa and salivary and mammary glands. Tonsils and mesenteric lymph nodes exhibit values intermediate between those of central and peripheral lymphoid systems. In patients with plasma IgA-deficiency, IgA 2 is the predominant intestinal IgA plasma cells. This may explain the frequent association of an asymptomatic condition and plasma IgA deficiency.     

Significance of different J chain profiles in human tissues: generation of IgA and IgM with binding site for secretory component is related to the J chain expressing capacity of the total local immunocyte population, including IgG and IgD producing cells, and depends on the clinical state of the tissue

Most IgA producing cells in normal intestinal and nasal mucosa synthesize dimers or larger polymers as evidenced by 90% cytoplasmic affinity for secretory component (SC) in vitro and almost 100% J chain positivity. The comparable median figures for normal exocrine glands (salivary, lacrimal, lactating mammary) were 84% and 92%, respectively. Conversely, IgA immunocytes in the subepithelial areas of palatine tonsils and in other extraglandular tissues, such as inflamed gingiva and intestinal submucosa, showed only 16-28% SC binding capacity and 18-51% J chain positivity. Similarly decreasing J chain expression, from glandular to extraglandular sites, was revealed not only for IgM immunocytes but also for those producing IgD or IgG, particularly the latter. This observation indicated more extensive overall clonal maturation in tissues without glandular elements since J chain expression seems to be a feature of relatively early memory B cells. The results were supported by studies in patients with selective IgA deficiency. Inflammatory disease caused significantly reduced SC binding capacity of IgA cells, both in intestinal mucosa and tonsils; this change was paralleled by decreased J chain expression, not only for mucosal and tonsillar IgA cells but also for mucosal IgG cells, suggesting local appearance of more mature clones. The resulting change to production of monomeric IgA may adversely affect secretory immunity and thus contribute to perpetuation of chronic inflammatory disease.     

Transforming growth factor α expression in normal gastric mucosa, intestinal metaplasia, dysplasia and gastric carcinoma—an immunohistochemical study

Using a monoclonal antibody (GF10) and a standard immunohistochemical technique, immuno-expression of transforming growth factor alpha (TGF-alpha) was found consistently within the differentiated compartment of normal adult human gastric mucosa. In 70% of mucosal samples exhibiting intestinal metaplasia there was more or less uniform TGF-alpha immunoreactivity throughout the intestinalized mucosa. Similarly, 60% of cases of dysplasia and 60% of gastric carcinomas showed strong immunoreactivity in most of the cells. However, whereas 93% of intestinal-type cancers showed strong immunoreactivity only 30% of the diffuse type were stained and then only weakly. Transforming growth factor alpha expression is thus a fairly regular feature of several types of differentiated gastric epithelia (normal, metaplastic, dysplastic and intestinal-type carcinoma), while its relative absence may be a factor in the histogenesis of the diffuse type of gastric carcinoma.     

In vitro synthesis of immunoglobulins, secretory component, complement and lysozyme by human gastrointestinal tissues. I. Normal tissues.

An in vitro culture technique has been used to demonstrate synthesis of proteins by human gastrointestinal tissues cultured in vitro. Histologically normal tissues were obtained endoscopically and surgically. IgA and secretory component (SC) were produced in all sites, but the relative intensity of IgA synthesis and SC synthesis varied. In stomach and small intestine the intensity of IgA synthesis was greater than that of SC, but in large bowel mucosa, there appeared to be an excess of SC synthesis. Synthesis of IgG and IgM was also found in all sites. Complement proteins were produced by some of the intestinal biopsies, and by parotid gland. Lysozyme was synthesized by parotid gland and by gastric mucosa, and to a lesser extent in small intestine, and rarely in large intestine. The results suggest that in addition to the local mucosal IgA system the local production of other immunoglobulins, as well as non-immunoglobulin humoral defence factors, may be important host defences of the normal gastrointestinal tract.     

Limited expression of APRIL and its receptors prior to intestinal IgA plasma cell development during human infancy

The absence of immunoglobulin A (IgA) in the intestinal tract renders young infants highly susceptible to enteric infections. However, mediators of initial IgA induction in this population are undefined. We determined the temporal acquisition of plasma cells by isotype and expression of T cell-independent (TI) and -dependent (TD) IgA class switch factors in the human intestinal tract during early infancy. We found that IgA plasma cells were largely absent in the infant intestine until after 1 month of age, approaching adult densities later in infancy than both IgM and IgG. The restricted development of IgA plasma cells in the first month was accompanied by reduced expression of the TI factor a proliferation-inducing ligand (APRIL) and its receptors TACI (transmembrane activator and calcium-modulator and cyclophilin ligand interactor) and B cell maturation antigen (BCMA) within isolated lymphoid follicles (ILFs). Moreover, both APRIL and BCMA expression strongly correlated with increasing IgA plasma cell densities over time. Conversely, TD mediators (CD40 ligand (CD40L) and CD40) were expressed within ILFs before 1 month and were not associated with IgA plasma cell generation. In addition, preterm infants had lower densities of IgA plasma cells and reduced APRIL expression compared with full-term infants. Thus, blunted TI responses may contribute to the delayed induction of intestinal IgA during early human infancy.     

Identification of Paneth cells in pyloric glands associated with gastric and intestinal mixed-type intestinal metaplasia of the human stomach

We have proposed that intestinal metaplasia (IM) of the human stomach be divided into two types on the basis of cell differentiation status: a gastric and intestinal (GI) mixed type and a solely intestinal (I) type. In the GI mixed type, gastric (foveolar epithelial and pyloric gland cells) and intestinal (goblet, intestinal absorptive, and Paneth cells) phenotype cells coexist in the same intestinalized gastric glands in various combinations and degrees. Consequently, intestinalized gastric glands are hybrids. Although we have described the rare appearance of Paneth-like cells in pyloric glands of GI mixed-type IM, the absence of an appropriate Paneth cell marker leaves room for doubt as to their true character. The purpose of this study was to clearly identify Paneth cells in pyloric glands in IM lesions using a new Paneth cell marker, a polyclonal antibody human defensin (HD)-5, raised against HD-5, which is included in granules of Paneth cells. A total of 105 gastric samples (4 biopsy and 101 surgical resected specimens) were examined. In only nine cases (8.6%), the antibody allowed demonstration of Paneth cells in pyloric glands in GI mixed-type IM, confirming our previous finding. Analysis of the proliferative cell (P) zone indicated that a common stem cell might generate both GI phenotype cells by upward and downward migration. No Paneth cells were found above the P zone. The results suggest that the stem cells show abnormal cell differentiation in IM lesions but preserve their normal direction of migration.     

Lymphoid follicles in antral mucosa: immune response to Campylobacter pylori?

The prevalence of lymphoid follicles in endoscopic biopsy specimens from normal antral mucosa (n = 220), mucosa with reflux gastritis (n = 104), and in cases with Campylobacter pylori-associated gastritis (n = 2544) was studied. In the latter group whether there were associations between degree and activity of gastritis and the prevalence of lymphoid follicles and between the occurrence of lymphoid follicles and the presence of intestinal metaplasia in the antrum were investigated. In cases with normal mucosa and in those with reflux gastritis lymphoid follicles were not detected, but mucosal lymphoid follicles were found in 1297 (54%) of the cases with C pylori-associated gastritis. The prevalence of lymphoid follicles in the antral mucosa depended on the degree and activity of the gastritis and also correlated with the presence of intestinal metaplasia. The development of lymphoid follicles in the mucosa of the antrum probably represents, primarily, an immune response to the colonisation of the mucosa by C pylori.     

IgE-containing cells in gastric mucosa with and without Helicobacter pylori infection

Gastric mucosa responds with inflammation to Helicobacter pylori (H. pylori) infection. While numerous reports have shown that the immune system produces specific IgG, IgA, and IgM isotype anti H. pylori antibodies, IgE-mediated pathways of H. pylori-associated gastritis are mostly unknown. Our aim was to evaluate whether an increased presence of IgE in the antral gastric mucosa is responsible for the severity of the H. pylori-associated gastritis. The number of IgE-containing cells was estimated in formalin-fixed, paraffin-embedded antral gastric biopsy specimens using immunohistochemistry in three groups of patients: (i) 20 H. pylori-positive cases with moderate inflammation, (ii) 19 H. pylori-negative cases with moderate inflammation, and (iii) 19 H. pylori-negative cases with normal mucosa. In chronic gastritis, the number of IgE-positive cells increased significantly as compared to normal mucosa. In gastritic patients, H. pylori positivity was accompanied by a significant accumulation of IgE-positive cells, mainly plasma cells. These data suggest that IgE-mediated immune response probably plays an important role in the development of H. pylori-associated gastritis.     

Apoptosis in human gastric mucosa, chronic gastritis, dysplasia and carcinoma: analysis by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling

We examined the existence and distribution of apoptotic cells in human gastric mucosa, chronic gastritis, adenomatous dysplasias and carcinomas in 15 surgically removed stomachs in which dysplasia and carcinoma were found simultaneously. Serial sections were cut for immunohistochemistry for p53 oncoprotein and Ki-67 antigen, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labelling (TUNEL). TUNEL signal-positive apoptotic cells were rare in normal mucosa, while a few apoptotic cells were noted in gastritic mucosa and intestinal metaplasia, intermingled with Ki-67 antigen-positive cells forming a generative cell zone. This suggests the cell-cycle-dependent apoptosis of gastric mucosa. The frequency of apoptotic cells per crypt was higher in incomplete than in complete metaplasia, implying greater underlying DNA damage in the former. TUNEL indices (TI; percentage of TUNEL-positive cells in tumour cells) were slightly higher in adenomatous dysplasias (4.9±2.1) than in carcinoma (3.9±1.1), but there was no no statistical difference. Ki-67 indices (KI; percentage of Ki-67 antigen-positive cells in tumour cells) were significantly (P<0.05) higher in carcinomas than in dysplasias. Thus, gastric adenomatous dysplasias were characterized by relatively higher TI and lower KI, which might reflect a more static growth potential. The expression of p53 oncoprotein in cancer cells is thought to be an apoptosis-suppressing event, although its precise role remains to be elucidated. Overall, these results indicate that apoptosis plays a crucial part in the morphogenesis of gastritic mucosa including intestinal metaplasia, and that the process is correlated both with tumourigenesis and with proliferative activity.     

Histological studies on the ontogeny of bovine gut-associated lymphoid tissue: appearance of T cells and development of IgG+ and IgA+ cells in lymphoid follicles

The development and distribution of lymphocyte subsets in bovine gut-associated lymphoid tissues (ileal and jejunal Peyer's patches (PP)) were examined. Before birth, the composition of lymphocyte subsets in both PP follicles did not differ except for the dimensions of the interfollicular area and the dome region. Many IgM+ cells were observed in these follicles, but very few CD3+, IgG+, and IgA+ cells could be found. At neonatal period, the IgG+ cells, which did not produce IgG mRNA, were dominant within both PP follicles. From 1 month after birth, many CD3+ cells, IgG mRNA expression, and IgA mRNA expression were detected within the jejunal PP follicles, but very few were in the ileal PP follicles. These data suggest that the characteristics of the jejunal PP follicles metamorphose into secondary lymphoid tissue such as germinal centers at around 1 month after birth, whereas the characteristics of ileal PP follicles were distinct from those of germinal centers.     

Immunohistochemical study of the lymph-follicles and lymphocytes in the gastric mucosa]

Immunohistochemical stainings according to ABC method (UCHL-1,L-26,MT-1,MB-1,IgG,IgA,IgD,IgM, kappa, lambda, LN-1 and LN-2) for the lymphocytes in the germinal center, mantle zone and infiltrative lymphocytes in the gastric mucosa of 30 cases of chronic gastritis and 10 cases of reactive lymphoreticular hyperplasia (RLH) were performed. Lymphocytes in the germinal center and mantle zone consisted usually of B-cells positively stained by L-26 and MB-1. However, in the interstitially infiltrative cells,T-cells positively stained by UCHL-1 and MT-1 were not infrequently contained. Immunoglobulin stainings revealed marked positivity for IgG,IgA,IgD,IgM, kappa and lambda in the interstitial lymphocytes and plasma cells. As to the RLH, small number of T-cells scattered in the germinal center and surrounding area of lymph follicles, and large number of T-cells were found among the follicles, where B-cells were more infrequently found than in the interstitial area of propria mucosae. Confusion of enlarged germinal centers and monotonous proliferation of lymphocytes among the lymph follicles showing monotonous positivity for the stains of heavy and light chains in the cases of RLH were suggestive of malignant change.     

Induction of Salivary Antibody Responses in Rats after Immunization in Peyer''s Patches

We examined salivary, milk, and serum antibody levels after immunization in the Peyer's patches (Pp) of rats with horse spleen ferritin. Priming of the Pp one day after parturition led to the appearance of IgG, but not IgA or IgM, anti-ferritin antibodies in saliva 9 days later. IgG and IgM antibodies were detected both in milk and in serum, whereas IgA antibodies could only be demonstrated in milk. During a second lactation period the salivary antibodies had vanished but IgG antibodies could still be detected in milk and serum. During a third lactation period, when the rats were immunized in the Pp a second time, not only IgG but also IgA anti-ferritin antibodies appeared in the saliva. Salivary IgG antibody levels and milk IgG, IgM, and IgA antibody levels were higher than those observed after primary immunization in the Pp. The IgG antibody activity in the saliva was positively correlated to the serum IgG antibody activity. It is concluded that salivary IgA antibody responses can be induced by immunization in the Pp. The results of this study suggests that IgA antibodies detected in saliva are produced locally by cells that have migrated from the intestinal lymphoid tissue to the salivary glands.     

Immunoglobulin-containing cells and the origin of immunoglobulins in the respiratory tract of sheep

The amounts of immunoglobulins formed in the respiratory tract of sheep were investigated by comparing the distribution of radiolabelled immunoglobulin between plasma and lymph from the caudal mediastinal lymph node efferent duct which drains 50-70% of lung lymph. In addition, immunoglobulin-containing cells in the lymph node lymph and in the respiratory mucosa were counted by immunofluorescence. Whereas 91.6 +/- 5.92% and 67.73 +/- 10.07% of IgG1 and IgG2 respectively were plasma-derived, only 35.89 +/- 7.09% of IgA was plasma-derived. IgM and albumin were wholly plasma-derived. There was no evidence for selective transport of any immunoglobulins from plasma into lymph. It was estimated that mediastinal lymph node lymph contributed 0.12 g IgA to the circulation daily, the bulk of this being of local origin. There were only few cells expressing IgA in the caudal mediastinal lymph node and its efferent lymph indicating that, unlike gut-associated lymphoid tissue, bronchus-associated lymphoid tissue in sheep is not a major site of IgA cell precursor production. Despite this finding, IgA was the most frequent isotype of Ig-expressing cells in the respiratory mucosa and it is concluded that the locally formed IgA in lymph node lymph originated from mucosal plasma cells. IgM- and IgG-expressing cells were much less numerous than IgA-cells in the mucosa, but were the predominant isotype in the caudal mediastinal lymph node and regional lymph nodes and in mediastinal lymph node efferent lymph. These experiments have established that the respiratory tract of sheep is qualitatively similar to the intestine with respect to immunoglobulin synthesis in that the bulk of IgA is locally derived and a smaller but significant local contribution of IgG1 and IgG2 occurs, but that it is a poor source of IgA cell precursors. This implies that the IgA plasma cell population in the respiratory mucosa probably originates from distant mucosal sites.     

Six types of intraepithelial vacuoles in the human gastric mucosa

Two types of goblet cells are classically described in the literature as markers of intestinal or colonic metaplasia of the gastric mucosa. In addition to those, four types of vacuolated cells in the gastric mucosa are herein described. One type corresponded to mucus-positive goblet cells (as in classical intestinal metaplasia) but containing in addition neuroendocrine granules. Another type presented vacuolated cells with cilia. Many of the ciliated cells had negative mucus reaction. Ciliated metaplastic cells were observed in cystically dilated pyloric glands. Another type was characterized by vacuolated cells without cilia, also in pyloric glands. The latter vacuoles often adopted an infranuclear position. Finally, cells with intraepithelial vacuoles (mucus-negative) centered by a lymphocyte were recorded in foveolar cells. Possible explanations for the occurrence of the type of intraepithelial vacuoles herein described were discussed. The occurrence of various types of intraepithelial vacuoles in the gastric mucosa should be borne in mind in the histological differential diagnosis of intestinal metaplasia in the gastric mucosa in H & E routine examinations.