Ultrastructural changes in gastric epithelium caused by bile salt

We assessed the influence of bile salt in neutral solution on the morphology and function of the Ussing-chambered stomach mucosa. Guinea pig gastric mucosa was exposed to 5, 10, and 20 mM taurocholic acid in balanced salt solution with glucose. Control tissues were exposed to salt solution without taurocholic acid. During exposure to the bile salt, the pH of the bathing solutions was maintained at 7.0-7.4 with 0.1 M sodium hydroxide. Potential difference, short-circuit current, and resistance of the tissues were recorded every 15 min. The morphologic effects of the bile salt were determined with light and electron microscopy. The 10 and 20 mM concentrations caused significant deterioration of the electrophysiologic measurements and major morphologic alteration on both light and electron microscopic evaluations. The 5 mM concentration of taurocholic acid did not affect the electrophysiologic measurements or the light microscopic appearance of the surface cells. Electron microscopy, however, revealed morphologic changes in over 50% of the specimens examined. These findings demonstrate that in vitro, mammalian gastric mucosa shows significant ultrastructural changes when exposed to physiological concentrations of bile salt in a neutral solution. These changes appear to be subtle in that they are evident on transmission electron microscopy even before significant change can be demonstrated by light microscopy or by standard electrophysiologic measurements.     

Gastric mucosal defense mechanisms: effects of salicylate and histamine.

Some of the recent concepts about the gastric mucosal defense mechanisms against damage by luminal acid and the effects of histamine and salicylate on these mechanisms are reviewed. The mucosal barrier to acid appears to consist of at least two physiologic components: a permeability mechanism and a metabolic mechanism related to cellular bicarbonate production as a result of acid secretion. In the absence of salicylate, histamine appears to exert some protection by affecting both mechanisms, but in the presence of salicylate, histamine's protective effect is limited to altering mucosal permeability. The actions of salicylate on the gastric mucosa are complex, related in part to the concentration of salicylate and the pH of the luminal fluid. The damaging effects of salicylate appear to be related more to the concentration of acid in the lumen than to the lipid solubility of the drug. Salicylate increases permeability regardless of pH; the increase is initially selective for cations and subsequently becomes nonselective, involving both cations and anions. Although both low and high concentrations of salicylate increase mucosal permeability to hydrogen ions, only high concentrations of salicylate affect cellular bicarbonate production.     

Duodenogastric reflux of bile acids, gastrin and parietal cells, and gastric acid secretion before and 6 months after cholecystectomy

In order to evaluate the effect of cholecystectomy on the gastric mucosa, the duodenogastric reflux of total and single bile acids, the number of parietal and gastrin cells, and the volume of gastric acid secretion were examined in 15 patients with gallstones and functioning gallbladders before and 6 months after cholecystectomy. The duodenogastric reflux of the total bile acids increased from a mean preoperative value of 1.9 mumol/hour to a mean postoperative value of 21 mumol/hour (p = 0.008). The duodenogastric reflux of all single bile acids increased after cholecystectomy, with a higher increase in glycoconjugated compared with tauroconjugated bile acids. The parietal cells decreased from a mean preoperative value of 82.8 to a mean postoperative value of 68.7 (p = 0.05), whereas there was only a mild increase in the number of gastrin cells; the output of gastric acid remained unchanged. The variation of the gastrin cells before and after cholecystectomy was negatively correlated only with the variation of taurocholic acid (r = -0.50, p = 0.05), while the variation of the parietal cells was mildly correlated with all single bile acids (r = 0.35-0.50, 0.05 less than p less than 0.02). These findings show an increased duodenogastric reflux of bile acids 6 months after cholecystectomy with a mild morphologic alteration of the gastric mucosa.     

Salicylate enhances rat gastric gelatinase activity

BACKGROUND: Increased matrix metalloproteinase (MMP) activity is associated with tissue injury in some organs. Their role in gut injury remains to be fully elucidated. We recently demonstrated that increased MMP-2 activity participated in lipopolysaccharide (LPS)-induced gastric injury. Thus we hypothesized that MMPs may play a role in other models of gastric injury. MATERIALS AND METHODS: The effect of L-NAME (10 mg/kg IP) or salicylate (100 mg/kg IP) on gastric injury from 20% ethanol was evaluated in an anesthetized model of gastric injury. In a separate experiment, gastric metalloproteinase activity was assessed after salicylate or L-NAME administration. Rats were given either L-NAME (10 mg/kg), salicylate (100 mg/kg), or saline IP and sacrificed after 6 hours. Gastric mucosa was harvested and portions of the glandular stomach snap frozen for gelatin and in situ zymography as indices of MMP activity. Subsequently the effect of MMP inhibition on macroscopic gastric injury from salicylate and a dilute luminal irritant was determined. RESULTS: Both L-NAME and salicylate significantly increased gastric injury from 20% ethanol versus saline controls. Salicylate treatment significantly increased gelatinase activity as determined by in situ zymography and gelatin zymography while L-NAME did not. MMP inhibition ameliorated macroscopic gastric injury secondary to salicylate and a dilute luminal irritant. CONCLUSIONS: This is the first study to report that MMP activity increases in the stomach following salicylate treatment. These data suggest that MMPs may play a role in the ability of salicylate to exacerbate gastric injury from irritants, but likely do not play a role in mediating the deleterious effects of L-NAME.     

Isolation and viability of gastric mucosal surface cells of the rabbit

This study describes a mechanical method for obtaining small sheets of gastric mucosal surface cells and an enzymatic digestion method for obtaining isolated gastric mucosal surface cells. Exclusion of vital dyes, maintenance of intracellular K⁺ and Na⁺ gradients, and O₂ consumption studies suggest that both preparations are viable. Histochemical stains indicate that both preparations are relatively pure. A major difference between the scraped and isolated surface cells is the rate of O₂ consumption. The response of the two preparations to glucose and to dinitrophenol, which interferes with mitochondrial respiration, suggests that the differences in oxygen consumption are not due to major alterations in oxidative phosphorylation. This correlates with electron microscopic examinations which show that mitochondrial structure does not appear to have been greatly affected by the isolation procedures. Alteration of the enzyme digestion method permitted isolation of gastric glandular cells. Expected differences in oxygen consumption and enzyme activity were observed between the isolated surface and glandular cell preparations.     

Long-term effects of bile on the gastric mucosa of the dog

Bile was allowed to act on the gastric mucosa of dogs for periods of time ranging from 60 to 540 days. Then the mucosa was studied by light microscopy and morphometric techniques to determine height of the mucosa, infundibulum and glands, number of mast cells, and number of interepithelial lymphocytes. Marked acute inflammatory lesions and decrease in cellular mucin content were observed mainly in the antral mucosa exposed to bile for up to 130 days. Other changes appearing both in the antrum and fundus of the same dogs included an increase in the number of interepithelial lymphocytes, architectural distortion, and decrease in the height of the mucosa, the infundibulum being the most affected component in the antrum and the glands in the fundus. Positive alkaline phosphatase activity was observed in the antral area up to 130 days. With continuous exposure to bile, the gastric mucosa appeared to undergo adaptation, since all the described alterations gradually disappeared.     

Bile salt binding by maalox, sucralfate, and meciadanol: in vitro and clinical comparisons

Inhibition of gastric acid secretion is a major factor in protecting the gastric mucosa, although other mechanisms such as bile salt binding may contribute to the protective properties of individual agents. Sucralfate, antacid (Maalox), and Meciadanol, a new flavonoid, were compared with cholestyramine resin for binding bile salts. The free, glycine, and taurine conjugates of the human bile salts, cholate, chenodeoxycholate, and deoxycholate, were incubated with each of the above. Cholestyramine resin adsorbed 91-97% of all bile salts tested. Meciadanol adsorbed all of the bile salts fairly well except for the free forms of chenodeoxycholate and deoxycholate. Meciadanol (53 to 84%) adsorbed bile salts better than sucralfate (4.2 to 61%), and significantly (P less than 0.05) better than Maalox (10 to 47%). In our in vitro studies, sucralfate was not as effective in binding bile salts as previously reported. Patients in the surgical intensive care unit were randomized prospectively to receive nasogastric instillation of Maalox, sucralfate, or Meciadanol to prevent gastrointestinal bleeding. The gastric aspirates were analyzed for bile salt concentration. The mean bile salt concentration of those treated with Maalox (0.24 mM), Meciadanol (0.24 mM), or sucralfate (0.35 mM) was significantly lower than those treated with nasogastric aspiration (0.87 mM) alone (P less than 0.01). This suggests that these substances bind bile salts and may provide additional protection to the gastric mucosa along with their ability to neutralize gastric acid.     

Regurgitant bile acids and mucosal injury of the gastric remnant after partial gastrectomy

Three groups, each consisting of seven patients who had undergone either Billroth I, Billroth II, or pylorus-preserving gastrectomies, were evaluated more than 18 months postoperatively in terms of concentration and amount of bile acids in the gastric aspirate and histologic changes in the gastric remnant mucosa. Concentrations of bile acids were determined by gas chromatography and mucosal specimens were obtained by endoscopic biopsy. The total bile acid concentration and all of the individual fractional bile acid levels, whether free or conjugated, were significantly higher in the Billroth II group than in the other two groups. The amount of gastric aspirate was also highest in the Billroth II group. Endoscopic biopsy revealed glandular dysplasia to be predominantly in the Billroth II group. The presence of bile acids in the gastric remnant may contribute to mucosal injury, possibly leading to cancer in the gastric remnant, especially after the Billroth II operation.     

Effect of melatonin on burn-induced gastric mucosal injury in rats

We studied the effect of melatonin treatment on gastric mucosal damage induced by experimental burns and its possible relation to changes in gastric lipid peroxidation status. Melatonin was intraperitoneally applied immediately after third-degree burns over 30% of total body skin surface area of rats. Malondialdehyde (MDA), uric acid (UA) and sulphydril (SH) levels were determined in gastric mucosa and blood plasma and used as biomarkers of the oxidative stress. The results showed that the skin burn caused oxidative stress evidenced by accumulation of MDA and UA as well as the depletion of SHs in gastric mucosa. Plasma MDA concentrations were elevated, while plasma SH concentrations were decreased after burns. Melatonin (10 mg per kg body weight) protected gastric mucosa from oxidative damage by suppressing lipid peroxidation and activating the antioxidant defence. It may be hypothesised that melatonin restores the redox balance in the gastric mucosa and protects it from burn-induced oxidative injury. Melatonin has no significant influence on the concentrations of plasma MDA and antioxidants after burn; therefore, it should largely be considered as a limiting factor for tissue-damage.     

Effect of obstructive jaundice on the electrophysiological characteristics of the gastric mucosa and gastric acid secretion in rats

To elucidate the effect of jaundice on the electrophysiological characteristics of the gastric mucosa and gastric acid secretion, gastric mucosal potential difference (PD) and gastric acid secretion were measured in rats with obstructive jaundice. Also transepithelial potential difference (TEPD), short circuit current (Isc) and transepithelial electrical resistance (Rt) were measured in the isolated gastric mucosa of rats with obstructive jaundice. Secondly, to confirm whether the alteration of these parameters were induced by jaundice and increased serum bile acids in the jaundiced rats, the effects of biliary drainage on the electrophysiological characteristics and gastric acid secretion, and the effects of bile acid (TCA) on TEPD, Isc, Rt were evaluated. PD, TEPD, Isc and gastric acid secretion were reduced in the jaundiced rats, and tended to recover after biliary drainage. TEPD and Isc were reduced significantly by TCA administration. These results suggest that active ion transport in the gastric mucosal cells and gastric acid secretion are impaired in jaundiced rats and the increased serum bile acid in jaundiced rats may cause these dysfunctions and the impaired active ionic transport function is improved by biliary drainage.     

Salicylate action on medullary inspiratory neuron activity in a brainstem-spinal cord preparation from newborn rats

Salicylate affects central respiratory control. The inspiratory neurons are the most important component of the medullary respiratory control center because they modulate the final motor output via the phrenic nerve. We investigated changes in burst rate, intraburst firing frequency, and membrane properties of inspiratory neurons in the isolated brainstem after the administration of salicylate. Newborn rat brainstem-spinal cord preparations were superfused with salicylate. Whole-cell recordings were performed from inspiratory neurons. Application of 1 mM salicylate caused an increase in the inspiratory neuronal burst rate from 6.9 +/- 1.6 bursts/min to 8.2 +/- 1.9 bursts/min (P < 0.05). The inspiratory neuron burst rate decreased from 8.3 +/- 0.7 bursts/min to 4.5 +/- 1.1 bursts/min after the application of 10 mM salicylate (P < 0.01). The depressant effect of 10 mM salicylate was antagonized by the gamma-aminobutyric acid (GABA) receptor antagonist bicuculline (1 microM). Resting membrane potential and intraburst firing frequency did not change with the application of salicylate and bicuculline even when the burst rate did change. We conclude that the effects of salicylate on the medullary inspiratory neurons are mainly due to a presynaptic action. GABAergic mechanisms are probably involved in the salicylate-induced central respiratory depression.     

Heterotopic gastric mucosa in the hilar bile duct mimicking hilar cholangiocarcinoma: report of a case

We herein report a case of heterotopic gastric mucosa in the hilar bile duct. An asymptomatic 58-year-old male was noted to have mild liver dysfunction in March 2009 during the follow-up for angina pectoris. Abdominal-enhanced CT revealed wall thickening from the upper common hepatic bile duct to the left hepatic bile duct. Endoscopic retrograde cholangiopancreatography (ERCP) showed stenosis at the junction of the left hepatic bile duct. Although the patient??s serum tumor markers were all within the normal ranges, the possibility of malignant disease of the biliary tree could not be ruled out. Left hepatectomy with the caudate lobe and resection of the extrahepatic bile duct were performed. Histopathologically, the resected specimen showed a polypoid lesion measuring 2 × 2 cm in size that projected into the lumen of the left hepatic bile duct. Microscopic examination revealed this polypoid lesion to be composed of mucous glands resembling gastric fundic glands, with parietal and chief cells. We also review eight other reports of heterotopic gastric mucosa in the biliary tree previously published in the English literature.     

Acidification of arterial blood enhances gastric mucosal injury induced by bile salts in dogs

The influence of intraarterial infusion of hydrochloric acid on gastric mucosal injury induced by topical bile salts was assessed in a canine ex vivo model. Exposure of the gastric mucosa to 5.0 mM sodium taurocholate in acid bathing solution alone resulted in a slight but not significant reduction in intramural pH of gastric mucosa and mild mucosal damage. When hydrochloric acid (0.1 N) was infused directly into the artery perfusing the stomach of the dogs, the intramural pH of gastric mucosa markedly decreased. The mucosal acidification was associated with severe ulcers in all test dogs that received intraarterial infusion of hydrochloric acid. The deleterious effect of intraarterial infusion of hydrochloric acid was not due to ischemia since the gastric blood flow remained relatively unchanged. The severe mucosal ulcerations were perhaps caused by the decrease in the ability of gastric blood to buffer the incoming luminal acid induced by topical bile salts. These results suggest that (1) the pH of arterial blood perfusing the stomach may also be an important determinant in the ability of the gastric mucosa to protect itself against acid injury, and (2) better management of systemic acid-base balance may contribute to a lower incidence of stress ulcers in critically ill patients.     

An evaluation of naloxone as a gastric cytoprotective agent during hemorrhagic shock

Administration of naloxone, an opiate antagonist, is known to improve survival from hemorrhagic shock and to reverse the effects of septicemia on gastric mucosal O2 tension and potential difference (PD). We tested these potentially cytoprotective actions in the ex-vivo canine gastric chamber model with acid, bile, and hemorrhagic hypotension. Naloxone (2 mg/kg IV bolus, then 2 mg/kg/hr IV) was given before or during shock. Naloxone did not affect oxygen consumption, the bile-induced drop in PD, or the transmucosal oxygen consumption, the bile-induced drop in PD, or the transmucosal movements of H+, Na+, K+, and fluid. The reduction in average mucosal lesion formation with naloxone pretreatment (5.4 +/- 1.2 vs. 2.8 +/- 0.5%) was not statistically significant (p = 0.07). Similarly, administration of naloxone after the onset of shock also failed to protect the mucosa from stress ulceration. We conclude: 1) naloxone does not inhibit the effects of topical bile on the gastric mucosal barrier; 2) naloxone has no apparent effect on local gastric vascular resistance during hemorrhagic shock; and 3) the therapeutic potential of naloxone as an anti-ulcer drug is questionable. 24GM 23095     

Decreased parietal cell acid secretion after vagotomy is not associated with altered gastric prostaglandin levels

In a previous investigation we demonstrated that after vagotomy there is a decreased ability of parietal cells to use intracellular cyclic adenosine monophosphate (cAMP). Prostaglandins are present in gastric mucosa and have been demonstrated to be inhibitors of in vivo and in vitro acid secretion through a cAMP-mediated mechanism. In the present study we have examined in vitro acid secretion and prostaglandin E2 levels in rabbits 8 weeks after vagotomy and pyloroplasty compared with control animals to investigate the possible role of prostaglandins in postvagotomy-impaired cAMP use. In vitro acid secretion was assessed in isolated gastric glands by 14C-labeled aminopyrine uptake and prostaglandin E2-generating capacity measured by high-pressure liquid chromatography. After vagotomy, there was a decrease in basal aminopyrine uptake (p less than 0.05), as well as that simulated by histamine and 8-bromo-cAMP (p less than 0.007). No differences were observed in prostaglandin E2 levels in either gastric glands or intact fundic mucosa (p greater than 0.5). These data suggest that impaired cAMP use observed in parietal cells after vagotomy is not the result of alterations in gastric prostaglandin levels.     

Alteration of gastric surface cell pH regulation by sodium taurocholate

Gastric mucosal cells from guinea pigs were grown in cell culture. Acridine orange fluorescence at 624 nm was used as an qualitative indicator of intracellular pH. The cultured cells were exposed to Hank's solution at pH 7.4, 6.0, 5.0, 4.0, 3.0, and 1.8 for 30 min. After removal of the acid, the cells were loaded with acridine orange for qualitative pH assessment. The cells developed alkaline shifts in the cytoplasmic pH in direct proportion to the acid load. This alkaline overcorrection after exposure to acid was blocked by amiloride and blunted by taurocholic acid in a dose-dependent fashion. These results suggest that gastric surface cells may regulate their cytoplasmic pH with a sodium-hydrogen antiporter. These results also suggest that this antiport system may be adversely affected by bile salts.     

Salicylate reduces cisplatin nephrotoxicity by inhibition of tumor necrosis factor-alpha

BACKGROUND: Salicylate was recently shown to provide protection against cisplatin nephrotoxicity in rats. We have demonstrated that enhanced tumor necrosis factor-alpha (TNF-alpha) production mediates, in part, cisplatin nephrotoxicity. The purpose of this study was to determine if the protective effects of salicylate were mediated through inhibition of TNF-alpha in vivo and to explore the mechanism of inhibition in vitro. METHODS: The effects of treatment with cisplatin alone and in combination with sodium salicylate in mice on renal function, histology, and gene expression were determined. The effects of cisplatin and salicylate on TNF-alpha expression, nuclear factor kappa B (NF-kappa B) activity, and apoptosis were determined in vitro using cultured murine proximal tubule cells. RESULTS: Salicylate significantly reduced both the functional and histologic evidence of cisplatin renal injury. Cisplatin increased the renal expression of TNF-alpha, monocyte chemoattractant protein-1 (MCP-1), intercellular adhesion molecule (ICAM)-1, heme oxygenase-1, TNF receptor 1 (TNFR1), and TNF receptor 2 (TNFR2). Treatment with sodium salicylate blunted the increase in TNF-alpha mRNA and also reduced serum TNF-alpha protein levels. Salicylate had little protective effect when administered with cisplatin to TNF-alpha-deficient mice. Cisplatin increased the degradation of I kappa B (I kappa B) in a time-dependent manner and also increased nuclear NF-kappa B binding activity. Salicylate inhibited I kappa B degradation and NF-kappa B binding activity in the presence of cisplatin. In addition, salicylate inhibited the cisplatin induced TNF-alpha mRNA increase in mouse proximal tubule epithelial (TKPT) cells. CONCLUSION: These results indicate that salicylate acts via inhibition of TNF-alpha production to reduce cisplatin nephrotoxicity. The inhibition of TNF-alpha production may be mediated via stabilization of I kappa B.     

The effect of reflux of bile juice on the development of residual stomach cancer

An experimental study was performed using an organ culture method to evaluate the effect of a duodenal juice reflux on the development of cancer in the residual stomach. The following results were as follows. An intracellular DNA levels to combine with carcinogenic agents was significantly increased in the mucosa of the residual stomach compared to the parietal mucosa in the whole stomach (control group). In the human gastric mucosa exposed to the bile acid, the intracellular DNA level to combined with carcinogenic agents was increased, and thus the effect of the bile acid as a surfactant on the experimental development of gastric cancer was suggested. An atrophic change was main feature of the residual stomach. Autoradiographic findings revealed that the proliferative zone was extended and a number of immature cells appeared which became to be target cells. Therefore, the residual stomach might provide a situation where the cancer would easily develop.     

Gastric stress ulcer of the rat: relative contribution of the pyloric sphincter, HCO3- bile reflux and mucosal blood flow

Gastric ulceration has been induced after stress, combining 24 h of fasting and 48 h of restraint in 9 groups of 20 rats with or without a pyloroplasty or a pylorojejunostomy combined with atropine and gastric infusion of NaHCO3 or taurocholic acid. After death or sacrifice at 48 h, ulcer index and blood in the jejunum were determined. Gastric mucosal blood flow was measured semi-continuously by a laser Doppler velocimeter. There were 45% deaths after 48 h of restraint alone, and 70% in the group combining pylorojejunostomy with taurocholic acid. Mortality was lower (p less than or equal to 0.01) pylorojejunostomy alone and more significantly so (p less than or equal to 0.001) when associated with NaHCO3. There was no death when NaHCO3 and atropine were combined with restraint. The mucosal blood flow increased significantly during the first 12 h of restraint in the taurocholic acid group. Both groups with NaHCO3 had mucosal blood flows similar to the controls. Gastric acid and gastric emptying, mucosal ischemia and bile reflux are joint factors inducing gastric stress ulcer. The 100% survival and the low ulcer index after a treatment by atropine and gastric infusion of NaHCO3 suggest that these well-known drugs should be used more frequently.