Ileal epithelial cell migration after 40% small-bowel resection

Adaptation in the small bowel after resection is associated with increases in crypt cell proliferation and villus height. This paper gives the results of an autoradiographic investigation with [³H]thymidine of epithelial cell migration 60 days after 40% small-bowel resection in the rat. The mean number of cell positions between the crypt-villus junction and the leading labeled cell 30 hr after injection was increased by 19.4% in the resected group (P<0.02). The mean total number of cells per villus column was increased by 27.8% (P<0.002). Migration rate estimated in cell positions per hour was accelerated by 18.9% (P<0.001) after resection. The 8.1% lengthened life span of villus cells was not statistically significant. The increased number of cells per villus column and unaltered life span of villus cells would facilitate functional adaptation. The causal relationship between the larger villus cell population and accelerated migration after resection and increased crypt cell proliferation is unknown.This investigation was supported by the National Health and Medical Research Council.     

Genetic ablation of parietal cells in transgenic mice: a new model for analyzing cell lineage relationships in the gastric mucosa.

The gastric mucosa of mammalian stomach contains several differentiated cell types specialized for the secretion of acid, digestive enzymes, mucus, and hormones. Understanding whether each of these cell lineages is derived from a common stem cell has been a challenging problem. We have used a genetic approach to analyze the ontogeny of progenitor cells within mouse stomach. Herpes simplex virus 1 thymidine kinase was targeted to parietal cells within the gastric mucosa of transgenic mice, and parietal cells were ablated by treatment of animals with the antiherpetic drug ganciclovir. Ganciclovir treatment produced complete ablation of parietal cells, dissolution of gastric glands, and loss of chief and mucus-producing cells. Termination of drug treatment led to the reemergence of all major gastric epithelial cell types and restoration of glandular architecture. Our results imply the existence of a pluripotent stem cell for the gastric mucosa. Parietal cell ablation should provide a model for analyzing cell lineage relationships within the stomach as well as mechanisms underlying gastric injury and repair.     

Omeprazole does not cause unscheduled DNA synthesis in rabbit parietal cells in vitro.

Parietal cells from rabbit gastric mucosa, enriched to greater than 90% purity, were used to study the effect of the H+,K(+)-ATPase inhibitor omeprazole on DNA in vitro. In this preparation, omeprazole undergoes acid-catalyzed conversion to its active form, the sulfenamide, which subsequently binds to luminal SH groups of the H+,K(+)-ATPase and thereby inhibits acid secretion. In the parietal cell fraction the S-phase inhibitor hydroxyurea (HU) decreased [3H]thymidine uptake by 40% as measured by liquid scintillation counting (LSC), presumably due to inhibition of scheduled DNA synthesis in contaminating stem cells. In the presence of HU, irradiation with ultraviolet light (UV) or treatment with the gastric carcinogen, 1-methyl-3-nitro-1-nitrosoguanidine (MNNG) increased [3H]thymidine uptake by a factor of 5. Autoradiography of isolated, stimulated parietal cells showed that UV irradiation and MNNG treatment increased the average number of silver grains over the nuclei 18-fold and 4-fold, respectively. In contrast, treatment of histamine-stimulated parietal cells with omeprazole or ranitidine in concentrations 100 times the IC50 value for inhibition of acid secretion in the parietal cells did not increase [3H]thymidine incorporation above the control levels, measured either by LSC or by autoradiography. Extracted DNA from stimulated parietal cells treated with [3H]omeprazole or [3H]MNNG showed no binding of [3H]omeprazole but considerable binding of [3H]MNNG. It is concluded that parietal cells can undergo DNA repair, but there is no indication that omeprazole, or its acid-derived metabolites, should cause any damage to DNA, nor does it bind to DNA in its target cell, where the highest concentrations of omeprazole and its acid-derived products are found.     

Cell kinetics of slow renewing cell populations in mice stomach

BACKGROUND: The renewal rates of parietal and chief cells in the gastric mucosa and smooth muscle cells of muscularis propria have not been examined as precisely as superficial epithelial cells. To examine cell renewal of these cells, continuous labeling with tritiated ([3H])-thymidine was performed. METHODS: Mice received 112 repeated injections of [3H]-thymidine at 6-hour intervals for 28 days after birth and were killed immediately thereafter, or 60, 120, 200 or 300 days after the last injection. RESULTS: After continuous labeling, most cells in the stomach were labeled. At 60 days, unlabeled parietal cells in the neck area of the gland and unlabeled chief cells in the middle part of the gland appeared. Thereafter, the area of unlabeled cells expanded downwards to the bottom of the gland. Times required for labeling of total cell populations of parietal and chief cells to half were less than 60 days and more than 200 days, respectively. At 300 days, most parietal cells and about half of the chief cells remained labeled in the bottom of the gland. The labeling index of smooth muscle cells was about 100% for 300 days. CONCLUSIONS: The time required for the newly formed parietal and chief cells to reach the lower end of the gland was more than 300 days. As a total cell population, the renewal rate of parietal cells was more rapid than that of chief cells. However, in terms of the downward migrating cell population, the renewal rate of parietal cells was a little slower than that of chief cells. Smooth muscle cells showed almost no renewal.     

Pyrimidine biosynthetic enzymes in rat intestine after small bowel resection.

After small bowel resection in the rat, mucosal hyperplasia and an increase in nucleic acid synthesis and cell proliferation occur in remaining small intestine. Male Sprague-Dawley rate underwent resection of 50 cm of proximal or distal intestine or sham operation. One month and 6 months after surgery, aspartate transcarbamylase, dihydroorotase, and uridine kinase were assayed in whole mucosa, and in some instances, in crypt mucosa ffrom the remaining intestinal segment. In control bowel, enzyme activity was significantly greater proximal compared with distal segments. One month after proximal or distal resection, mucosal enzyme activity per cm of gut was greater in the remnant bowel compared with controls. There was no such difference at 6 months. Specific enzyme activity of whole mucosa did not increase after resection because the assay was influenced by the disproportionately large contribution of villous protein. Specific enzyme activity (including thymidine kinase) of isolated crypt mucosa was significantly increased 1 month after distal resection. In addition, [3H]thymidine uptake into DNA of crypt mucosa from proximal remnants was also significantly increased. These results indicate that after small bowel resection, the enzymes of pryimidine biosynthesis increase in remaining bowel and parallel the accelerated rate of cell proliferation.     

Tritiated thymidine radioautographic study on the origin and renewal of secretin cells in the rat duodenum

The origin and renewal of secretin cells in the duodenum were investigated using the unlabeled antibody peroxidase-antiperoxidase technique and radioautography in rats killed at various times after single or multiple injections of [3H]thymidine. Secretin cells were spatially distributed from the upper crypt to the villus tip, being particularly numerous in the upper two-thirds of the duodenal villi. After a single injection of [3H]thymidine, there were no labeled secretin cells, indicating a lack of self-replicating activity. After repeated injections of the isotope, labeled secretin cells appeared and increased in number. They first occurred at the upper part of the crypt and the lower part of the villus, and later at the villus tip. All these cells were found to be labeled after continuous labeling for 120 h, which is considered to be the renewal time for this cell population.     

Radioautographic and quantitative studies on parietal and peptic cell kinetics in the mouse. A selective effect of gastrin on parietal cell proliferation.

Gastrin was given to mice subcutaneously at 8-hr intervals for 20 days. Control animals received only saline injections. With use of [3H]thymidine and radioautography, a significant increase in the production of new parietal cells (P less than 0.01) was observed in the gastrin-treated group. Increases in fundic progenitor cell DNA synthesis and shortened maturation time of the latter were demonstrated. Both could explain the observed changes in parietal cell kinetics. The quantitative estimation of parietal cell population after the administration of gastrin showed an increase in average parietal cell count per unit (P less than 0.01) and in total parietal cell number (P less than 0.05). This was consistent with the observed acceleration in the rate of parietal cell production under influence of gastrin. In contrast, no radioautographic difference in the proliferative activity of the peptic cells was observed between the gastrin-treated and the control mice. Moreover, at the end of the experiment, no changes in average peptic cell count per unit area, or in total peptic cell number, occurred under the effect of gastrin. The observation that peptic cells are renewed in a different way, independently from fundic progenitor cells and from parietal cells, was consistent with their peculiar lack in proliferative response to the hormone.     

Effects of parenteral hydrocortisone sodium succinate on epithelial renewal in hamster gastric mucosa

The aim of this study was to determine whether parenteral administration of steroids affects epithelial renewal in hamster stomach. Male golden hamsters received either hydrocortisone sodium succinate or saline intraperitoneally for three days. In the first experiment, hamsters were sacrificed 1 hr after injection of tritiated thymidine [( 3H]TdR) to label proliferating cells. In the second experiment, hamsters were sacrificed hourly after a single [3H]TdR injection up to 48 hr in order to determine cell cycle time by the method of fraction of labeled mitoses. In the third experiment, hamsters were sacrificed 1, 24, and 72 hr after [3H]TdR injection for the study of epithelial migration and cell turnover time. Sections of fundic and antral mucosae were prepared for light autoradiography. Steroid treatment caused no gross or microscopic injury to gastric mucosa, but the number of [3H]TdR-labeled cells as well as the thickness of the proliferative zone were reduced significantly in fundic mucosa, but not in antral mucosa. The study of the fraction labeled mitoses indicated that steroid treatment lengthened the cell cycle time in fundic mucosa, which was due primarily to prolonged G1 and DNA synthesis phases. Furthermore, epithelial migration was significantly slower in fundic mucosa after steroid treatment, which was associated with a prolonged cell turnover time. Thus, parenteral steroids depress the entire process of epithelial renewal in hamster fundic mucosa.     

FSH stimulation of DNA synthesis in Sertoli cells in culture.

The incorporation of [2H]thymidine into nuclear DNA was investigated in cultured Sertoli cells prepared from testes of 20-day-old rats. Addition of follicle-stimulating hormone (FSH) or dibutyryl cyclic, 3',5'-adenosine monophosphate (DBCAMP) to the culture medium greatly increased incorporation, expressed either as total amounts of [3H]thymidine incorporated per mug DNA or as the percentage of Sertoli cells with labeled nuclear DNA. No stimulation was observed in cells cultured in the presence of testosterone, insulin or cyclic 3',5'-GMP (cGMP). Light and electron microscopic autoradiographic analysis was employed to establish the identity of Sertoli cells having labeled nuclear DNA. Contaminating spermatogonia, which also took up labeled [3H]thymidine, were excluded from cell counts. In addition, Sertoli cells prepared from testes of irradiated 20-day-old germinal cell depleted rats were also observed to incorporate more [3H]thymidine into nuclear DNA when cultured in a chemically defined medium in the presence of FSH. DNA synthesis was abolished by prior treatment of cells with cytosine arabinoside. In separate experiments, the incorporation of [3H]thymidine into DNA of peritubular myoid cells was shown to be independent of FSH or dbcAMP.     

Increased follicular heterogeneity in experimental colloid goiter produced by refeeding iodine excess after thyroid hyperplasia

Delayed morphological changes induced in mouse hyperplastic thyroid by refeeding iodine were analyzed by light and electron microscopy, stereology, and autoradiography. Thyroid hyperplasia was induced by a low iodine diet supplemented with 0.25% propylthiouracil for 10 days. Involution was obtained by discontinuing the propylthiouracil and returning either to a moderate iodine diet [(MID) 1 microgram I/day] or to an iodine-rich diet [(HID) 10 micrograms I/day] for 40 days. In other experiments, three cycles of hyperplasia (8 days) and subsequent involution (8 days) with MID or HID were brought about. Control animals were fed MID or HID. All animals were killed when 12-14 weeks old after injection of 10-50 microCi 125I. Double labeling, with repeated injections of [3H]thymidine from day 0 to day 7 of involution followed by 125I injection 4 h before killing, was also performed. When involutions were performed with MID, most morphological variables returned to control values. However, when involution was brought about with HID, the glandular weight, the number of follicles, and the relative volume of follicular lumina remained larger than in controls. Moreover, the 125I-labeling pattern of the follicles was altered. The proportions of unlabeled, and unevenly or partly labeled, follicles, which were fewer than 5% in control groups, represented 25-35% of all follicles after involution with HID, whereas they were unchanged with MID. In unlabeled follicles the epithelium was flattened, with a reduced number of microvilli. Partly labeled follicles were of two types. In some follicles a persistent ring reaction was observed, suggesting an abnormally slow mixing of thyroglobulin. In others, the 125I labeling was restricted to areas adjacent to the apex of a reduced number of cells, suggesting that some cells were iodinating thyroglobulin, whereas others were not. There was no relationship between the follicular 125I labeling and the frequency of [3H]thymidine-labeled cells. These results indicate that refeeding iodine excess after hyperplasia leads to the formation of a colloid goiter with new follicles, and to an increased heterogeneity of iodine metabolism among follicles and among cells.     

Dominance of the Senescent Phenotype in Heterokaryons Between Replicative and Post-Replicative Human Fibroblast-Like Cells

In heterokaryons between senescent and young diploid fibroblast-like cells, dominance of the former with respect to nuclear DNA synthesis (incorporation of [³H]thymidine) was demonstrated. For identification of the respective partners, double-layer autoradiography was used after the old cells were labeled with [³H]methionine and the young cells were labeled with [¹⁴C]thymidine. Synchrony of nuclear labeling (i.e., all nuclei in a cell labeled with [³H]thymidine) was observed in the majority of di- and polykaryons during the second and third of three 24-hr periods of labeling with [³H]thymidine. The results are compatible with either terminal differentiation or error theories of clonal senescence.     

Kinetic studies on the development of the adult population of Leydig cells in testes of the pubertal rat

The objective of this study was to determine whether postnatal increases in rat Leydig cell number result from differentiation of precursor cells, division of existing Leydig cells, or both. Our approach was 1) to examine changes in the absolute number of Leydig cells and potential precursor cells (macrophages, pericytes, and mesenchymal, endothelial, and myoid cells) per testis on day 19 of gestation (day -2) and days 7, 14, 21, 28, and 56 postpartum; 2) to examine the frequency with which mesenchymal and Leydig cells divide during prenatal and postnatal development; and 3) to identify and examine the fate of the progeny of Leydig and mesenchymal cell divisions during prenatal and postnatal development. Stereological methods were used to show that mesenchymal cells comprised 44% of the total interstitial cell population and Leydig cells 16% on day -2, whereas by day 56 postpartum the relationship had reversed; mesenchymal cells comprised 3% and Leydig cells 49%. These results suggested a precursor-product relationship between mesenchymal and Leydig cells because no such reciprocal relationship was observed between Leydig cells and macrophages, pericytes, endothelial, or myoid cells. Autoradiographic analysis of [3H]thymidine incorporation into mesenchymal and Leydig cells was consistent with this interpretation. In a series of pulse-chase experiments, the percentage of labeled mesenchymal and Leydig cells was measured after a single injection of [3H]thymidine on days 2, 14, 28, and 56 postpartum, each followed by sampling at timed intervals (between 1 h and 14 days) thereafter. Starting on day 14, the percentage of labeled Leydig cells was approximately 1% immediately after injection of [3H]thymidine and increased significantly to approximately 6% by 6 days after injection. No such increase was observed when rats were similarly injected starting on days 2, 28, and 56 postpartum. The rise in Leydig cell labeling between days 14 and 28 postpartum did not result in a decline in the number of silver grains over labeled Leydig cell nuclei, indicating that the increase in the percentage of labeled cells was not caused by Leydig cell division. These observations led us to conclude that the increase in Leydig cell labeling from days 14 to 28 was the result of recruitment from a compartment of labeled mesenchymal cells. In contrast, our analysis indicated that from day 28 postpartum and thereafter until the mature number of Leydig cells is attained, Leydig cells are generated by division of morphologically recognizable Leydig cells.     

Effect of carbenoxolone on the synthesis of glycoproteins and DNA in rat gastric epithelial cells.

The influence of carbenoxolone on the synthesis of glycoproteins in the surface mucous cells and the production of new cells in the rat gastric mucosa was studied by means of a vascular perfusion system. The rate of incorporation of tritiated galactose, glucosamine, serine, and sulphate in surface mucous cells, studied by autoradiography, was not affected by the addition of carbenoxolone to the drinking water. The sugar composition (determined by gas-liquid chromatography) of the gastric glycoproteins (isolated by centrifugation in CsCl), was not changed in carbenoxolone-treated rats. Compared with untreated animals, the number of [3H]-thymidine labelled nuclei per fundic pit increased by 38% to 76% in carbenoxolone-treated rats, implying a higher number of mitotically active cells. This results in an increased supply of young mucous cells; if this also proves to be true in human gastric mucosa, it may be relevant to the therapeutic effect of carbenoxolone.     

Observations on a proposed measure of genotoxicity in rat gastric mucosa.

Current methods for the study of the toxicological effects of antisecretory medications on the gastric mucosa possess disadvantages or limitations. A novel assay has been proposed to assess gastric mucosal genotoxicity in which the proton-pump inhibitor omeprazole has been reported to induce direct damage to cellular DNA, raising questions about the safety of this drug. To define the applicability of this proposed measure of genotoxicity and to examine the effects of omeprazole in this assay, control agents, known carcinogens, and omeprazole in various doses and formulations were administered to rats by gavage, followed by [3H]thymidine labeling of DNA in vivo approximately 14 hours later. The incorporation of the [3H]thymidine label into DNA of gastric mucosal cells liberated by limited pronase digestion was in close agreement with published results for negative and positive controls. Omeprazole, administered in doses ranging from 10 mg/kg to 300 mg/kg, did not increase [3H]thymidine incorporation into cellular DNA in this assay. The gastric carcinogen 1-methyl-2-nitro-1-nitrosoguanidine at 20 and 50 mg/kg increased [3H]thymidine incorporation. Pretreatment in vivo with hydroxyurea before [3H]thymidine labeling to inhibit replicative DNA synthesis suppressed [3H]thymidine incorporation more than 97% in negative controls and MNNG and more than 93% in omeprazole treatments. This indicates that replicative DNA synthesis was almost totally responsible for the [3H]thymidine incorporation and the contribution of unscheduled DNA synthesis to the total [3H]thymidine incorporation is minor. Flow cytometric analysis of the cell cycle of the gastric mucosal cells liberated by the limited pronase digestion indicated significant contamination of the preparation with dividing cells (4% in negative controls and 14% in MNNG-treated positive controls). These findings indicate that the proposed screening assay for genotoxicity in rat gastric mucosa is not a reliable measure of unscheduled DNA synthesis in its present form, and conclusions about genotoxic effects of any drug using this assay as initially proposed appear questionable.     

Studies of secretory activity and pathomorphology of the gastric mucosa in taeniasis (Taenia saginata)

Studies were performed on 149 patients, aged 18 to 50, hospitalized in the Department of Parasitic and Tropical Diseases, Medical Academy in Poznań. The following examinations were performed: gastric contents testing with histamine stimulus according to Kay, the histological and cytological examination of gastric mucosa biopsies. Among patients with Taenia saginata invasion, disturbances in secretory activity of gastric mucosa were demonstrated in 57.7% of studied patients. Hypoacidity was found in 49.7% of patients; it appeared both in the oldest group, over 40 years (71.4%), as well as in two younger, although at two times lower frequency (38.2 and 39.6%, respectively). At set of histological and cytological studies was performed on gastric mucosa biopsies in a chosen group of 30 patients (average age of 26.2 years) with taeniasis and hypoacidity, before and after the treatment of the invasion. Before the treatment cellular infiltrates consisting of mononuclear cells were observed in gastric mucosa biopsies; cytological analysis of gastric gland cross-sections demonstrated 3 to 4 times lower number of parietal cells as compared to that of mucous cells. After the treatment of taeniasis, secretory activity of gastric mucosa returned to the normal level in 20 patients (66.7% of total). At the same time, the histological and cytological analysis of the biopsies showed decreased intensity of cell infiltrates in the glandular layer of the mucosa and an increased number of parietal cells as compared to the number of mucous cells. In 10 patients (33.3% of total), hypoacidity of the gastric juice as well as gastric mucosa lesions persisted despite the elimination of T. saginata. No relation could be found between the duration of T. saginata invasion on the one hand and secretory disturbances on the other. The subsidance of functional disturbances in gastric mucosa and of their morphological exponents in 2/3 of patients after taeniasis treatment indicates a causal relationship between time of taeniasis and secretory disturbances and histological lesions in gastric mucosa.     

Reversible drug-induced oxyntic atrophy in rats

BACKGROUND & AIMS: Oxyntic atrophy is the hallmark of chronic gastritis. Many studies have sought to develop animal models for oxyntic atrophy, but none of them are reversible. We now report that rats administered high doses of DMP 777 demonstrate reversible oxyntic atrophy. METHODS: DMP 777 was administered to CD-1 rats by oral gavage (200 mg. kg(-1). day(-1)). Serum gastrin level, in vivo acid secretion, and gastric histological changes were evaluated in DMP 777-dosed animals. Direct effects of DMP 777 on parietal cells were evaluated by assessment of aminopyrine accumulation into isolated rabbit parietal cells, as well as by assessment of DMP 777 effects on acridine orange fluorescence and H(+),K(+)-adenosine triphosphatase (ATPase) activity in isolated tubulovesicles. RESULTS: Oral dosing with DMP 777 caused a rapid increase in serum gastrin levels and severe hypochlorhydria. DMP 777 inhibited aminopyrine accumulation into rabbit parietal cells stimulated with either histamine or forskolin. DMP 777 reversed a stimulated proton gradient in isolated parietal cell tubulovesicles. Oral dosing with DMP 777 led to rapid loss of parietal cells from the gastric mucosa. In response to the acute loss of parietal cells, there was an increase in the activity of the progenitor zone along with rapid expansion of the foveolar cell compartment. DMP 777 treatment also led to the emergence of bromodeoxyuridine-labeled cells and cells positive for periodic acid-Schiff in the basal region of fundic glands. With extended dosing over 3-6 months, foveolar hyperplasia and oxyntic atrophy were sustained while chief cell, enterochromaffin-like cell, and somatostatin cell populations were decreased. No histological evidence of neoplastic transformation was observed with dosing up to 6 months. Withdrawal of the drug after 3 or 6 months of dosing led to complete restitution of the normal mucosal lineages within 3 months. CONCLUSIONS: DMP 777 acts as a protonophore with specificity for parietal cell acid-secretory membranes. DMP 777 in high doses leads to the specific loss of parietal cells. Foveolar hyperplasia, loss of normal gland lineages, and the emergence of basal mucous cells appear as sequelae of the absence of parietal cells. The results suggest that parietal cells are critical for the maintenance of the normal mucosal lineage repertoire.     

Prostaglandin prevents alterations in DNA,RNA, and protein in damaged gastric mucosa

Fasted rats were given either 16,16-dimethyl-PGE2 (dmPGE2) (1 microgram/kg) or normal saline subcutaneously followed by the oral administration of 1 ml of 100% ethanol or saline 30 min later. At 1, 3, 6, and 24 hr later, animals were sacrificed, their stomachs examined for necrotic ulcerations, and the incorporation of [3H]thymidine into DNA as well as tissue levels of DNA, RNA, and protein content of glandular mucosa determined. Compared with control animals, severe ulcerations of 70-80% of the glandular mucosa were observed in rats given 100% ethanol at all time periods. Accompanying these ulcerations were marked depressions in tissue levels of DNA and RNA at 1, 3, 6, and 24 hr after exposure to ethanol, and protein at 1, 3, and 6 hr following ethanol. In rats pretreated with dmPGE2 before ethanol administration, these alterations in tissue levels of DNA, RNA, and protein were prevented as were ulcerations of the glandular stomach at each time period. Synthesis of mucosal DNA was not significantly different from control rats in any of the groups studied. It is concluded that (1) gastric mucosal damage by alcohol is associated with a decrease in tissue levels of DNA, RNA, and protein; (2) dmPGE2 maintains normal tissue levels of DNA, RNA, and protein by preventing the shedding of mucosal cells by alcohol; and (3) the ability of dmPGE2 to prevent gastric damage by alcohol is not mediated through stimulation of DNA synthesis.     

Na+/HCO3- cotransport and expression of NBC1 and NBC2 in rabbit gastric parietal and mucous cells.

BACKGROUND & AIMS: The gastric epithelium protects itself against luminal acid by secreting HCO3--rich fluid into the mucous layer and by HCO3--dependent intracellular pH regulation, but the basolateral HCO3- uptake mechanisms are incompletely characterized. This study examined the expression and functional significance of the Na+/HCO3- cotransporters NBC1 and NBC2 in rabbit gastric epithelial cells. METHODS: Rabbit NBC1 and NBC2 complementary DNA fragments were cloned and sequenced, and cellular expression levels were assessed by semiquantitative polymerase chain reaction. Na+/HCO3- cotransport activity was measured fluorometrically in cultured rabbit parietal and mucous cells. RESULTS: NBC1 expression was 4.5-fold lower in the stomach than kidney cortex and 5.5-fold higher in mucous than parietal cells. NBC2 expression in the stomach was much lower than in the eye, approximately 4-fold lower than NBC1 expression in the stomach, and 2.5-fold higher in mucous than parietal cells. The Na+- and HCO3--dependent, dimethylamiloride-insensitive (which at 500 micromol/L completely inhibits all Na+/H+ exchanger isoforms) base influx rates were 4.6 +/- 0.02 and 16.2 +/- 0.04 mmol/L/min in acidified parietal and mucous cells, respectively, and were not significantly different in the absence of Cl-. CONCLUSIONS: This study shows that NBC1 and NBC2 are expressed in rabbit stomach, with high levels in mucous cells where Na+/HCO3- cotransport is the major base-importing mechanism in the presence of CO2/HCO3-.     

Abnormal pattern of cell proliferation in the entire colonic mucosa of patients with colon adenoma or cancer

Using autoradiography after 1 h of pulsed labeling with tritiated thymidine in endoscopic biopsy specimens from normal-appearing mucosa, cell proliferation was determined at six predetermined sites of the whole colon in patients with neoplastic disease of the large bowel and was compared with that of subjects without macroscopic colonic pathology. The labeling index (the percentage of cells incorporating [3H]thymidine) was 8.6 +/- 0.5 (mean +/- SEM) in 13 patients with colon carcinoma (p less than 0.001 vs. 16 control patients whose labeling index was 4.9 +/- 0.2) and 9.1 +/- 0.4 in 11 patients with a large adenoma in the colon (p less than 0.001 vs. controls). Twenty-one patients with one or more small adenomas (diameter less than 1 cm) had a moderately increased cell proliferation compared with controls (labeling index 6.2 +/- 0.3, p less than 0.02 vs. controls). In patients with neoplastic disease an enlargement of the proliferative compartment was found, whereas 6 patients with Crohn's colitis had values for labeling index and a distribution of labeled cells along the crypt comparable to that of control subjects. An increased cell proliferation was found along the entire colon under each of the neoplastic conditions studied. These findings indicate that although neoplastic lesions develop in a limited area of the colon, the entire large bowel may be at risk for tumor growth.     

A monolayer culture of human gastric epithelial cells

Our aim was to develop a fibroblast-free monolayer culture of human gastric mucosal cells, using the specimens obtained by routine endoscopic biopsy. Human gastric mucosa obtained from normal volunteers by endoscopic biopsy was dissociated from collagenase and hyaluronidase. Dissociated cells were cultured in supplemented Coon's modified Ham's F-12 medium. Within 24 hr of inoculation, the cells were attached to the culture dishes. This was followed by cellular outgrowth. On phase-contrast microscopy, all cells had epithelial characteristics and fibroblasts were not observed. Ninety percent of cells contained periodic acid Schiff reaction-positive mucous granules after diastase digestion consistent with mucous epithelial cells. Two percent of the cells gave a strong reaction for succinic dehydrogenase activity (parietal cells). Immunohistochemical staining for pepsinogen in cultured cells was negative. On EM, microvilli-like projections, junctional complexes, Golgi apparatus, and mucous granules were apparent in the majority of cells. Mitotic figures were observed by day 3 with Giemsa staining. Autoradiographically, these cells were able to incorporate [3H]TdR into the nuclei. Cells were capable of synthesizing DNA, and this function was inhibited by cycloheximide. Cells could be cultured for up to two weeks without fibroblast contamination. A method of primary monolayer culture of human gastric mucosa obtained by a routine endoscopic biopsy has been successfully developed.