CGRP modulates mucin synthesis in surface mucus cells of rat gastric oxyntic mucosa

We examined the effects of the calcitonin gene-related peptide (CGRP), including the possible participation of nitric oxide (NO), on mucin biosynthesis in the surface epithelium and remaining deep mucosa as well as the entire mucosa and compared the distribution of CGRP and NO synthase (NOS) using a combination of double immunofluorescence labeling and multiple dye filter. Pieces of tissue obtained from the corpus and antrum were incubated in a medium containing [(3)H]glucosamine and CGRP, with or without the NOS inhibitor. CGRP dose-dependently enhanced [(3)H]glucosamine incorporation into the corpus mucin but had no effect on antral mucin biosynthesis. The CGRP receptor antagonist, CGRP-(8-37), prevented the increase in (3)H-labeled corpus mucin. This stimulation of corpus mucin synthesis disappeared after removal of the surface mucus cell layer. CGRP activated the mucin biosynthesis in the surface mucus cells. In the full-thickness corpus mucosa, CGRP-induced activation was completely blocked by the NOS inhibitor. CGRP-immunoreactive fibers were intertwined within the surface mucus cell layer with type I NOS immunoreactivity. These results show that CGRP-stimulated mucin biosynthesis mediated by NO is limited to surface mucus cells of the rat gastric oxyntic mucosa.     

Stimulant effect of nitric oxide generator and roxatidine on mucin biosynthesis of rat gastric oxyntic mucosa.

Although the involvement of nitric oxide (NO) in an increasing gastric mucus metabolism has been reported, information on whether or not its activation is limited to the specific mucus-producing cells is lacking. In this paper, we report the effect of the exogenous NO-donor, isosorbide dinitrate (ISDN), and second-generation histamine H2 receptor antagonist roxatidine (2-acetoxy-N-(3-[m-(1-piperidinylmethyl)phenoxy]propyl)acetamide hydrochloride) which is demonstrated to accelerate the mucin metabolism mediated by endogenous NO, on the mucin biosynthesis in distinct sites and layers of the rat gastric mucosa using an organ culture technique. Radiolabeled mucin was obtained from the tissue of full-thickness and the deep corpus layer, and the antrum of the rat stomach incubated for 5 hr with [3H]glucosamine(GlcN) in vitro. With the addition of ISDN to the culture medium, 3H-labeled mucin in the full-thickness corpus mucosa increased to 124-145% of the control (p<0.05), but not in the antrum. This stimulation of the mucin synthesis disappeared by the removal treatment of the surface mucous cell layer which has immunoreactivity of neuronal NO synthase. Similarly, roxatidine stimulated the mucin biosynthesis in the full-thickness corpus mucosa, but not in the gland mucous cell layer. These results suggest that the stimulation of the mucin biosynthesis mediated by NO is restricted to the surface mucous cells of the rat gastric oxyntic mucosa.     

The gastric mucus layers: constituents and regulation of accumulation

The mucus layer continuously covering the gastric mucosa consists of a loosely adherent layer that can be easily removed by suction, leaving a firmly adherent mucus layer attached to the epithelium. These two layers exhibit different gastroprotective roles; therefore, individual regulation of thickness and mucin composition were studied. Mucus thickness was measured in vivo with micropipettes in anesthetized mice [isoflurane; C57BL/6, Muc1-/-, inducible nitric oxide synthase (iNOS)-/-, and neuronal NOS (nNOS)-/-] and rats (inactin) after surgical exposure of the gastric mucosa. The two mucus layers covering the gastric mucosa were differently regulated. Luminal administration of PGE(2) increased the thickness of both layers, whereas luminal NO stimulated only firmly adherent mucus accumulation. A new gastroprotective role for iNOS was indicated since iNOS-deficient mice had thinner firmly adherent mucus layers and a lower mucus accumulation rate, whereas nNOS did not appear to be involved in mucus secretion. Downregulation of gastric mucus accumulation was observed in Muc1-/- mice. Both the firmly and loosely adherent mucus layers consisted of Muc5ac mucins. In conclusion, this study showed that, even though both the two mucus layers covering the gastric mucosa consist of Muc5ac, they are differently regulated by luminal PGE(2) and NO. A new gastroprotective role for iNOS was indicated since iNOS-/- mice had a thinner firmly adherent mucus layer. In addition, a regulatory role of Muc1 was demonstrated since downregulation of gastric mucus accumulation was observed in Muc1-/- mice.     

Interactions of EGF and ornithine decarboxylase activity in the regulation of gastric mucus synthesis in cigarette smoke exposed rats

Cigarette smoking has been shown to aggravate ulceration and delay ulcer healing. Smokers had a lower level of mucus in their stomachs. In the present study, we examined whether cigarette smoke or its extract reduced mucus production through the suppression of epidermal growth factor (EGF) associated with the reduction of polyamine biosynthesis both in vivo and in vitro. Ornithine decarboxylase (ODC) activities and mucus synthesis were determined in rat gastric mucosa and in human MKN-28 cells. Incubation of MKN-28 cells with EGF (0.01-1.00 ng/mL) significantly increased mucus synthesis in vitro, which was accompanied by an increase of ODC activity. Removal of salivary glands decreased the circulated EGF level and induced a significant reduction of mucus-secreting layer thickness in the gastric mucosa. Cigarette smoke or its extract markedly decreased mucus synthesis in vivo and in vitro, both of which could be completely reversed by intravenous administration of EGF (20 microg/kg) in rats or co-incubation with EGF (1 and 2 ng/mL) in MKN-28 cells. However, ODC activities, which were suppressed by cigarette smoke or its extract, were unaffected by intravenous administration of EGF in rats, or only partially reversed by co-incubation with EGF in MKN-28 cells. These findings indicate that both EGF and ODC activity represent two different entities in the modulation of cigarette smoking on gastric mucus synthesis. The action of EGF on mucus synthesis may only be partially if not dependent on ODC activity in the stomach.     

Role of salivary epidermal growth factor in the maintenance of physicochemical characteristics of oral and gastric mucosal mucus coat

The involvement of salivary epidermal growth factor (EGF) in the maintenance of oral and gastric mucosal mucus coat dimension and chemical characteristics was investigated using sialoadenectomized rats. Examination of the oral and gastric mucosal surface by phase contrast microscopy and Alcian blue uptake revealed that deprivation of salivary EGF caused a 31-36% reduction in mucus coat thickness and a 38-43% reduction in adherent mucin content. Chemical analyses indicated that the mucus coat of sialoadenectomized group exhibited a 21-28% increase in protein and a 67% decrease in covalently bound fatty acids, a 30% decrease in carbohydrates, and a 32-37% decrease in lipids. Sialoadenectomy also evoked changes in the chemical composition of mucus glycoprotein component of oral and gastric mucus coat reflected in the lower content of sulfate (25-26%), associated lipids (24-25%), and covalently bound fatty acids (67-75%). Intragastric supplementation of EGF had no effect on the physicochemical changes caused by sialoadenectomy in the oral mucosal mucus coat, while nearly complete restoration to normal characteristics occurred in the gastric mucosal mucus coat. The results suggest that salivary EGF is essential for the maintenance of mucus coat dimension and quality needed in the protection of alimentary tract epithelium.     

The synthesis and secretion of gastric mucus glycoprotein by mucosal cells cultured in the presence of ethanol

The effect of ethanol on the synthesis and secretion of mucus glycoprotein in gastric mucosal cells was investigated. The mucosal cell suspensions were subjected to a short-term (4 h) culture in the presence of 0-1.5 M ethanol, with [3H]proline and [3H]palmitic acid as markers for glycoprotein synthesis and acylation. The synthesized labeled mucus glycoprotein was isolated from the incubation medium (extracellular glycoprotein) and from the mucosal cells (intracellular glycoprotein), and analyzed. Depending upon the ethanol concentration in the cell culture medium, two distinct effects on the synthesis and secretion of mucus glycoprotein were observed. The cells cultured in the presence of 0.02-0.1 M ethanol showed increased ability for the incorporation of [3H]proline and [3H]palmitic acid, and for the secretion of the newly assembled mucus glycoprotein. The synthesis of the glycoprotein increased 18-fold, acylation 5-fold, and secretion 10-fold. The synthesized glycoprotein, however, contained four to five times less of acyl-bound fatty acids. Ethanol at 0.1-1.5 M caused a marked reduction (62-64%) in the mucus glycoprotein synthesis, but the amount of glycoprotein released to the medium remained constant. This indicated that higher concentrations of ethanol caused the release of the preformed intracellular mucus glycoprotein reserves. The results demonstrate that gastric mucosal cells incubated in the presence of ethanol exhibit impaired synthesis and secretion of mucus glycoprotein, and that the severity of impairment depends upon the ethanol concentration.     

Biosynthesis, processing and secretion of mucus glycoprotein in the rat stomach

For the study of the biosynthesis, processing and secretion of mucus glycoproteins in rat gastric mucous cells, antibodies were raised against purified gastric mucus glycoproteins and against deglycosylated gastric mucus glycoproteins. Indirect immunofluorescence analysis of gastric mucosa sections revealed that both antibodies specifically labelled the mucus glycoprotein-synthesizing cells in the gastric mucosa. Stomach segments were pulse-labelled with [35S]cysteine and chased for various times. The radioactively labelled (glyco)proteins were quantitatively immunoprecipitated and analyzed by SDS-polyacrylamide gel electrophoresis. Less than 3% of the total radioactivity incorporated in protein was found to be present in mucus glycoproteins. Antibodies raised against native mucus glycoproteins recognized only high-molecular-weight mucus glycoproteins, while the antibodies against deglycosylated glycoproteins also bound to probable precursor forms. The synthesis of mature mucus glycoproteins (Mr greater than 300 000) required about 90 min. After 3 h of chase, only a small portion of the pulse-labelled mucus glycoproteins had been secreted; the majority of the radioactive glycoproteins at that time was still associated with the tissue. Immature (glyco)proteins were not secreted into the medium.     

Fast renewal of the distal colonic mucus layers by the surface goblet cells as measured by in vivo labeling of mucin glycoproteins

The enormous bacterial load and mechanical forces in colon create a special requirement for protection of the epithelium. In the distal colon, this problem is largely solved by separation of the bacteria from the epithelium by a firmly attached inner mucus layer. In addition, an outer mucus layer entraps bacteria to be cleared by distal transport. The mucus layers contain a network of Muc2 mucins as the main structural component. Here, the renewal rate of the inner protective mucus layer was studied as well as the production and secretion of Muc2 mucin in the distal colon. This was performed by intraperitoneal injection of N-azidoacetyl-galactosamine (GalNAz) that was in vivo incorporated during biosynthesis of O-glycosylated glycoproteins. The only gel-forming mucin produced in the colon is the Muc2 mucin and as it carries numerous O-glycans, the granulae of the goblet cells producing Muc2 mucin were intensely stained. The GalNAz-labeled glycoproteins were first observed in the Golgi apparatus of most cells. Goblet cells in the luminal surface epithelium had the fastest biosynthesis of Muc2 and secreted material already three hours after labeling. This secreted GalNAz-labeled Muc2 mucin formed the inner mucus layer. The goblet cells along the crypt epithelium accumulated labeled mucin vesicles for a longer period and secretion of labeled Muc2 mucin was first observed after 6 to 8 h. This study reveals a fast turnover (1 h) of the inner mucus layer in the distal colon mediated by goblet cells of the luminal surface epithelium.     

Decreased colonic mucus in rats with loperamide-induced constipation

Constipation is a risk factor of colorectal cancer. Mucin is a major component of lumenal mucus, which protects the colorectal mucosa against mechanical and chemical damage. The aim of this study was to evaluate mucus production and to quantitate lumen mucus in a rat model of spastic constipation. We induced constipation with loperamide (1.5 mg/kg), and histochemically evaluated mucus production and the thickness of the mucus layer at the fecal surface. We quantitated the mucus attached to the mucosal surface using colonic perfusion with N-acetylcysteine. While more feces remained in the colon, there was less fecal excretion and lower fecal water content in loperamide-administered rats than in control rats. Crypt epithelial cells contained less mucus in constipated rats than in control rats. The mucus layer at the fecal surface was thinner and less mucus was recovered from the mucosal surface in constipated rats than in control rats. Mucus production of crypt epithelial cells and mucus at the fecal and mucosal surface were reduced by loperamide-induced constipation.     

Luminal leptin activates mucin-secreting goblet cells in the large bowel

Leptin has been suggested to be involved in tissue injury and/or mucosal defence mechanisms. Here, we studied the effects of leptin on colonic mucus secretion and rat mucin 2 (rMuc2) expression. Wistar rats and ob/ob mice were used. Secretion of mucus was followed in vivo in the rat perfused colon model. Mucus secretion was quantified by ELISA, and rMuc2 mRNA levels were quantified by real-time RT PCR. The effects of leptin alone or in association with protein kinase C (PKC) and phosphatidylinositol 3-kinase (PI3K) inhibitors on mucin secreted by human mucus-secreting HT29-MTX cells were determined. Leptin was detected in the rat colonic lumen at substantial levels. Luminal perfusion of leptin stimulates mucus-secreting goblet cells in a dose-dependent manner in vivo in the rat. Leptin (10 nmol/l) increased mucus secretion by a factor of 3.5 and doubled rMuc2 mRNA levels in the colonic mucosa. There was no damage to mucosa 24 h after leptin, but the number of stained mucus cells significantly increased. Leptin-deficient ob/ob mice have abnormally dense mucus-filled goblet cells. In human colonic goblet-like HT29-MTX cells expressing leptin receptors, leptin increased mucin secretion by activating PKC- and PI3K-dependent pathways. This is the first demonstration that leptin, acting from the luminal side, controls the function of mucus-secreting goblet cells. Because the gel layer formed by mucus at the surface of the intestinal epithelium has a barrier function, our data may be relevant physiologically in defence mechanisms of the gastrointestinal tract.     

Vasoactive intestinal peptide stimulates mucus secretion, but nitric oxide has no effect on mucus secretion in the ferret trachea.

Vasoactive intestinal peptide (VIP) and nitric oxide (NO) are neurotransmitters involved in the regulation of bronchial and pulmonary vascular tone. Published studies of the effects of VIP on airway mucus secretion have yielded conflicting results. The purpose of this study was to determine the effect of VIP on mucus secretion in the ferret trachea and if this effect was influenced by NO. We used a sandwich enzyme-linked lectin assay to measure mucin secretion and a turbidimetric assay to measure lysozyme (serous cell) secretion from ferret tracheal segments. VIP (10(-7) M) increased mucin secretion over 2 h. VIP (10(-9) to 10(-5) M) stimulated mucin secretion in a dose-dependent fashion. VIP-induced mucin secretion was partially blocked by a VIP receptor antagonist (a chimeric VIP-pituitary adenylate cyclase-activating peptide analog, VIP receptor antagonist) at a 10-fold excess concentration. At all concentrations tested, neither NG-nitro-L-arginine methyl ester, an inhibitor of NO synthase, nor S-nitroso-N-acetyl-penicillamine, an NO donor, had any significant effect on constitutive or VIP-induced mucus secretion. We conclude that VIP-stimulated mucin and lysozyme secretion was both time dependent and dose dependent and that NO neither stimulates nor inhibits mucus secretion in the ferret trachea.     

In vitro acylation of rat gastric mucus glycoprotein with [3H]palmitic acid

The incorporation of fatty acids into gastric mucus glycoproteins was studied by incubating rat gastric mucosal cell suspensions with [9,10-3H]palmitic acid and [3H]proline. The mucus glycoprotein polymer, secreted into the growth medium (extracellular) and that contained within the cells (intracellular), was purified from the other components of the secretion, thoroughly delipidated, and then analyzed for the radiolabeled tracers. Both pools of mucus glycoprotein, incubated in the presence of [3H]palmitic acid, contained radioactive label which could not be removed by gel filtration, CsCl density gradient centrifugation, sodium dodecyl sulfate-gel electrophoresis, or lipid extraction. Treatment of the purified mucus glycoprotein with 1 M hydroxylamine or 0.3 M methanolic KOH released the radioactivity, thus indicating that [3H]palmitic acid was covalently bound by ester linkage to the glycoprotein. The released radioactivity was associated mainly (87%) with palmitic acid. The incorporation ratio of [3H]proline to [3H]palmitic acid was 0.12:1.0 in the extracellular glycoprotein and 1.38:1.0 in the intracellular glycoprotein, which suggested that acylation of mucus glycoprotein occurs in the intracellular compartment after completion of its polypeptide core. The fact that incorporation of [3H]palmitic acid was greater in the glycoprotein subunits than in the glycoprotein polymer indicates that acylation takes place near the end of subunit processing but before their assembly into the high molecular weight mucus glycoprotein polymer.     

Role of mucus in gastric mucosal protection.

Even though there is no general agreement as to the mechanism of gastric mucosal protection, the consensus is that the initial brunt of luminal insults falls on the mucus layer which constitutes the only identifiable physical barrier between the gastric lumen and the mucosal surface. The continuous renewal and resilient nature of this layer efficiently counters peptic erosion of the gel, assures its viscoelastic and permselective properties, and provides a milieu for containment of the diffusing luminal acid by mucosal bicarbonate. Disturbances in this delicate balance lead to the impairment of the protective function of mucus resulting in gastric disease. Indeed, the weakening of gastric mucosal defense is intimately associated with the diminished viscoelastic qualities of mucus, decrease in hydrogen ion retardation capacity, and the extensive proteolysis of its mucin component. Although until recently the disintegration of the mucus coat was attributed exclusively to the enhanced activity of intragastric pepsin, our studies provided strong argument that a bacterial factor, namely infection by Helicobacter pylori, through the action of its protease and lipase enzymes also is highly detrimental to the integrity of gastric mucus. Hence, agents capable of interfering with the pathogenic activity of this bacteria are becoming the drugs of choice in peptic ulcer therapy.     

应激状态下NO的胃粘膜保护作用及其与壁细胞泌酸的关系

目的:探讨应激状态下一氧化氮(NO)的胃粘膜保护作用及其与壁细胞泌酸的关系.方法:采用水浸-束缚应激(WRS)方法制备应激性溃疡(SU)动物模型,检测胃粘膜溃疡指数(UI)、胃粘膜NO含量和壁细胞H ,K -ATPase活性,观察L-硝基精氨酸甲酯(L-NAME)和L-精氨酸(L-Arg)对应激后大鼠壁细胞H ,K -ATPase活性及胃粘膜损伤的影响.结果:L-NAME(20 mg·kg-1)可使胃粘膜NO含量减少(P＜0.01),壁细胞H ,K -AT-Pase活性增加(P＜0.05),并加重应激所致的胃粘膜损伤;L-Arg(300 mg·kg-1)则使胃粘膜NO含量增加(P＜0.01),壁细胞H ,K -ATPase活性下降(P＜0.05),减轻应激所致胃粘膜损伤.结论:NO对应激状态下大鼠胃粘膜具有保护作用,其机制与抑制壁细胞H ,K -ATPase活性有关.     

Direct protective action of epidermal growth factor on isolated gastric mucosal surface epithelial cells.

Epidermal growth factor (EGF) protects gastric mucosa against acute injury produced by a variety of damaging agents, but the mechanism of its protective action is not clear. Since the surface epithelial cells (SEC) are important component of gastric mucosal defence, we studied whether EGF may directly protect isolated gastric SEC against ethanol injury in vitro, in condition independent of systemic factors and whether endogenous prostaglandins may play a role in EGF's protective action. The isolated SEC from rat gastric mucosa were preincubated in medium only, or medium containing 0.0001-10.0 micrograms/ml of h-rEGF for 15 minutes, and incubated with 8% ethanol for 1 hour. In another study the above experiment was repeated but cells were pretreated with 10(-4) or 10(-5) M indomethacin before EGF treatment. The cell viability was assessed by fast green exclusion test. Incubation of SEC with 8% ethanol significantly reduced SEC cell viability to 50 +/- 2%: EGF 0.1 or 1.0 microgram/ml significantly reduced ethanol induced damage (cell viability 59 +/- 3 and 62 +/- 3% respectively). Pretreatment with 10(-4) M indomethacin (the dose which does not affect SEC viability but inhibit PGE2 and PGI2 generation), significantly reduced protective action of EGF against 8% ethanol injury. EGF 1.0 and 10.0 micrograms/ml alone without ethanol increased PGE2 and 6 keto PGF1 alpha generation by SEC. These studies demonstrated: 1) EGF is able to protect gastric surface epithelial cells directly without mediation by systemic factors. 2) EGF induced protection of SEC may in part be mediated by prostaglandins.     

Co-localization of TFF2 with gland mucous cell mucin in gastric mucous cells and in extracellular mucous gel adherent to normal and damaged gastric mucosa

Trefoil factor 2 (TFF2) is mucin associated peptide that has a mucosal barrier function in addition to participating in repair and healing. We examined the localization of TFF2 and gastric mucins in gastric mucous cells, the surface mucous gel layer (SMGL) adherent to normal gastric mucosa, and in the mucoid cap covering gastric erosions. Carnoy’s solution, or formalin/picric acid-fixed paraffin embedded materials from resected stomachs and formalin-fixed paraffin embedded gastric biopsy materials were used. Sections were immunostained for the TFF2 and histochemically stained for gastric mucins. In addition, thick sectioned gastric mucosa fixed in Carnoy’s solution were stained with FITC-labeled GSA-II lectin specific for gland mucous cell mucin and examined for three-dimensional images of the SMGL using a confocal laser scanning microscope. The TFF2 and gland mucous cell mucin were found intermixed together in the gastric gland mucous cells, in the SMGL in laminated layers, and in the mucoid cap. A laminated arrangement of continuous sheets of gland mucous cell mucin in the SMGL was demonstrated in the three-dimensional images. Co-localization of the TFF2 with gland mucous cell mucin suggests a physical interaction between the TFF2 and gland mucous cell mucin. The TFF2 trapped in the adherent mucins may be responsible for mucosal defense, healing, and repair.     

Differences in potential and content of the mucus in the stomach of the rat with avitaminosis A

Vitamin A (vit. A) acts in the synthesis of glycoproteins and in cell surface phenomena of epithelia. Since the glycoproteins of gastric mucus and the integrity of gastric cell membranes are components of gastric barrier (GB), vit. A could play a role in GB. Five groups of rats were used: I) rats fed on vit. A deficient diet; II) rats pair-fed plus a daily oral dose of 45 micrograms vit. A; III) normal rats; IV) rats recovered from avitaminosis A (avit. A) after 20 days of daily oral dose of 300 micrograms vit. A; V) rats pair-fed plus a daily oral dose of 45 micrograms vit. A. We measured: 1) transparietal gastric potential difference (PD) in vivo (by means of agar-KCl electrodes); 2) mucus (by binding of Alcian blue): in gastric mucosa; adherent to gastric mucosa; in gastric lumen; 3) dry weight of the stomach. Avit. A induced: i) a decrease of PD and mucus in mucosa and lumen; ii) an increase of mucus adherent to mucosa; iii) an increase of the percentage of dry weight on wet weight. All parameters were normal after recovery from avit. A. Results suggest that avit. A could reduce either mucus synthesis or its erosion. Moreover avit. A might modify mucus structure and sterical configuration of mucosal cells. The alteration of mucosal cell membranes could decrease PD. In conclusion the modifications of some components of rat GB seem specifically caused by avit. A and suggest a protective role of vit. A.     

Essential components of antimicrobial gastrointestinal epithelial barrier: specific interaction of mucin with an integral apical membrane protein of gastric mucosa

BACKGROUND: The gastric mucosal protective barrier consists of two essential elements: mucus glycoprotein, mucin, secreted by gastric mucosal cells, and the mucin binding protein (MBP), an integral component of the apical epithelial membrane. The studies described here provide evidence on the structure of MBP, its interaction with mucin, and the susceptibility to phospholipase C (PLC) and Helicobacter pylori protease. MATERIAL AND METHODS: The rat gastric mucosa was used to isolate mucin and the apical epithelial membranes. A buffered saline extract of the mucosal cells was used for the isolation of mucin and the 1% Triton X-100-insoluble gastric apical membranes for the preparation of MBP. RESULTS: The studies on MBP, the mucosal mucin receptor revealed that the protein is anchored in apical membrane through glycosylphosphatidylinositol (GPI). The deamination of MBP with nitrous acid afforded phosphatidylinositols (PIs) and a water soluble, 97 kDa glycosylated protein. The in situ studies with untreated rat gastric mucosa and the mucosa depleted of mucin showed that MBP without mucin was susceptible to the proteolytic degradation with pepsin and H. pylori proteases, but was not released from the apical membrane by the treatment with bacterial PLC. CONCLUSION: The study of carbohydrate ligands for MBP revealed binding of octa- and decasaccharides of gastric mucin. The severe impairment in mucin adhesion to MBP, induced by the diet containing ethanol, supports the conclusion that specific carbohydrate determinants participate in mucin attachment to MBP and epigenetic control of the processes that coordinates its interaction with apical mucosal epithelium in the formation of innate protective barrier.     

Biosynthesis of gastric mucus glycoprotein of the rat

We have studied the biosynthesis of rat gastric mucin in stomach segments using an antiserum against rat gastric mucin specific for peptide epitopes. Pulse-chase experiments were performed with [35S]methionine, [3H]galactose, and [35S]sulfate to label mucin precursors in different stages of biosynthesis, which were analyzed after immunoprecipitation. The earliest mucin precursor that could be detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was a 300-kDa protein. The occurrence of N-linked "high-mannose" oligosaccharides on this protein was shown by susceptibility to degradation by endo-beta-N-acetylglucosaminidase H. This precursor could be labeled with [35S]methionine and not with [3H]galactose or [35S]sulfate. The 300-kDa precursor was converted into mature mucin after extensive glycosylation and sulfation. The mature mucin but not the 300-kDa precursor was in part secreted into the medium. Specific inhibition of sulfation with sodium chlorate had no effect on rate and amount of mucin secretion. In addition, we show that two core proteins are expressed in rats, slightly varying in Mr among individual animals.     

Effects of traditional herbal medicine on gastric mucin against ethanol-induced gastric injury in rats

The effect of the traditional herbal medicine, Rikkunshi-to and its component crude drugs, Zingiberis Rhizoma and Glycyrrhizae Radix, on the gastric mucin was studied using a method developed to separate and quantify the mucin localized in the different layers of rat gastric mucosa. The oral administration of spray-dried extract to Rikkunshi-to (1000 mg/kg), Zingiberis Rhizoma (500 mg/kg) and Glycyrrhizae Radix (500 mg/kg) significantly prevented gastric mucosal damage induced by 70% ethanol in rats. In ethanol-treated rats the mucin content of the deep mucosa was reduced, and the reduction of the deep corpus mucin content was significantly inhibited by pretreatment of Rikkunshi-to and Zingiberis Rhizoma. Rikkunshi-to and Glycyrrhizae Radix pretreatment increased the surface mucin content by 140 and 146%, respectively. The effect on the gastric mucin by each drug differed in the different layers of the gastric mucosa.